The purpose of this study was to analyze the effects of illite/smectite clay (I/S) on lignocellulosic degradation and humification process via metagenomics during cattle manure composting. The test group (TG) with 10% I/S and the reference group (RG) were established. The results indicated that the addition of I/S made the degradation rate of lignocellulose in TG (1.56%, 29.01%, 17.96%) was higher than that in RG (1.16%, 17.24%, 13.14%). Compared with RG, the abundance values of AA2, AA10, <em>GH1</em>, <em>GH1</em>0 in TG increased by 15.18%, 29.28%, 31.08%, 21.65%, respectively. Meanwhile, humic substance (HS) content was increased by 3.49% and 7.16% during RG and TG composting. Furthermore, the microbial community in TG changed, in which the relative abundance of Actinobacteria increased and the relative abundance of Proteobacteria decreased. Redundancy analysis (RDA) showed that the temperature was positively correlated with the abundance of the four enzymes, whereas the organic matter (OM) content was negatively correlated with them. Overall, adding I/S to the compost could stimulate microbial activity, promote the degradation of lignocellulose and humification process.

Keywords: Composting; Humification process; Illite/smectite clay; Lignocellulose breakdown; Metagenomics analysis.

The diversity and transcription efficiency of GH1 family β-glucosidase genes were investigated in natural and inoculated composts using a DNA clone library and real-time qPCR. Compositional differences were observed in the functional community between both composting processes. Proteobacteria, Actinobacteria, Firmicutes, and Chloroflexi were the dominant phyla. Twenty representative β-glucosidase genes were quantitatively analyzed from DNA and RNA pools. Principal component analysis and Pearson's correlation analysis showed that cellulose degradation is correlated with the composition and succession of functional microbial communities, and this correlation was mainly observed in Proteobacteria and Actinobacteria. Compared with inoculated compost, the functional microbial communities in natural compost with a low diversity index exhibited weak buffering capacity for function in response to environmental changes. This may explain the consistency and dysfunction of cellulose degradation and transcriptional regulation by dominant β-glucosidase genes. Except for the β-glucosidase genes encoding constitutive enzymes, individual β-glucosidase genes responded to environmental changes more drastically than the group β-glucosidase genes. Correlation results suggested that β-glucosidase genes belonging to Micrococcales played an important role in the regulation of intracellular β-glucosidase. These results indicated that the responses of functional microorganisms were different during both composting processes, and were reflected at both the individual and group levels.

Chromatin structure is critical for gene expression and many other cellular processes. In Arabidopsis thaliana, the floral repressor FLC adopts a self-loop chromatin structure via bridging of its flanking regions. This local gene loop is necessary for active FLC expression. However, the molecular mechanism underlying the formation of this class of gene loops is unknown. Here, we report the characterization of a group of linker histone-like proteins, named the GH1-HMGA family in Arabidopsis, which act as chromatin architecture modulators. We demonstrate that these family members redundantly promote the floral transition through the repression of FLC A genome-wide study revealed that this family preferentially binds to the 5' and 3' ends of gene bodies. The loss of this binding increases FLC expression by stabilizing the FLC 5' to 3' gene looping. Our study provides mechanistic insights into how a family of evolutionarily conserved proteins regulates the formation of local gene loops.

The current differential diagnosis for a short child with low insulin-like growth factor I (IGF-I) and a normal growth hormone (GH) peak in a GH stimulation test (GHST), after exclusion of acquired causes, includes the following disorders: (1) a decreased spontaneous GH secretion in contrast to a normal stimulated GH peak ("GH neurosecretory dysfunction," GHND) and (2) genetic conditions with a normal GH sensitivity (e.g., pathogenic variants of GH1 or GHSR) and (3) GH insensitivity (GHI). We present a critical appraisal of the concept of GHND and the role of 12- or 24-h GH profiles in the selection of children for GH treatment. The mean 24-h GH concentration in healthy children overlaps with that in those with GH deficiency, indicating that the previously proposed cutoff limit (3.0-3.2 μg/L) is too high. The main advantage of performing a GH profile is that it prevents about 20% of false-positive test results of the GHST, while it also detects a low spontaneous GH secretion in children who would be considered GH sufficient based on a stimulation test. However, due to a considerable burden for patients and the health budget, GH profiles are only used in few centres. Regarding genetic causes, there is good evidence of the existence of Kowarski syndrome (due to GH1 variants) but less on the role of GHSR variants. Several genetic causes of (partial) GHI are known (GHR, STAT5B, STAT3, IGF1, IGFALS defects, and Noonan and 3M syndromes), some responding positively to GH therapy. In the final section, we speculate on hypothetical causes.

Keywords: GH1; GHSR; Growth hormone; Growth hormone receptor; IGF1R; IGFALS; Insulin-like growth factor I; STAT5B; Short stature.

Most of the presently known β-glucosidases are sensitive to end-product inhibition by glucose, restricting their potential use in many industrial applications. Identification of novel glucose tolerant β-glucosidase can prove a pivotal solution to eliminate end-product inhibition and enhance the overall lignocellulosic saccharification process. In this study, a novel gene encoding β-glucosidase BglNB11 of 1405bp was identified in the genome of Saccharomonospora sp. NB11 and was successfully cloned and heterologously expressed in E. coli BL21 (DE3).The presence of conserved amino acids; NEPW and TENG indicated that BglNB11 belonged to GH1 β-glucosidases. The recombinant enzyme was purified using a Ni-NTA column, with the molecular mass of 51 kDa, using SDS-PAGE analysis. BglNB11 showed optimum activity at 40 °C and pH 7 and did not require any tested co-factors for activation. The kinetic values, K_m, V_max, k_cat, and k_cat/K_m of purified enzyme were 0.4037 mM, 5735.8 μmol/min/mg, 5042.16 s^-1 and 12487.71 s^-1 mM^-1, respectively. The enzyme was not inhibited by glucose to a concentration of 4 M but was slightly stimulated in the presence of glucose. Molecular docking of BglNB11 with glucose suggested that the relative binding position of glucose in the active site channel might be responsible for modulating end product tolerance and stimulation. β-glucosidase from BglNB11 is an excellent enzyme with high catalytic efficiency and enhanced glucose tolerance compared to many known glucose tolerant β-glucosidases. These unique properties of BglNB11 make it a prime candidate to be utilized in many biotechnological applications.

Keywords: Glucose tolerant; Molecular docking; Novel β-glucosidase; Saccharomonospora sp. NB11.

The control of the biological rhythms begins with the activation of photo- and thermosensitive cells located in various organs of the fish such as brain, eye, and skin, but a central clock is still to be identified in teleosts. Thermal changes are stressors which increase cortisol and affect the rhythm of other hormones such as melatonin and growth hormone (GH), in both endo- and ectothermic organisms. Our aim was to investigate how temperature (23°C for 6 days) lower than the optimal (28°C) modulates expression of several gene pathways including growth hormone (gh1) and its receptors (ghra, ghrb), insulin-like growth factor1 (igf1a, igf1b) and its receptors (igf1ra, igf1rb), cortisol and its receptor (gr), the limiting enzyme of melatonin synthesis (arylalkylamine N-acetyltransferase, aanat) and melatonin receptors (mtnr1aa, mtnr1bb), as well as their relationship with clock genes in Danio rerio in early light and early dark phases of the day. Lower temperature reduced the expression of the hormone gene gh1, and of the related receptors ghra, ghrb, igf1ra, and igf1rb. Cortisol levels were higher at the lower temperature, with a decrease of its receptor (gr) transcripts in the liver. Interestingly, we found higher levels of aanat transcripts in the brain at 23°C. Overall, lower temperature downregulated the transcription of hormone related genes and clock genes. The results suggest a strong correlation of temperature challenge with the clock molecular mechanism and the endocrine systems analyzed, especially the growth hormone and melatonin axes, in D. rerio tissues.

Keywords: Danio rerio; clock genes; hormone receptors; hormones; temperature.

Until recently, four types of cellobiose-fermenting Saccharomyces cerevisiae strains have been developed by introduction of a cellobiose metabolic pathway based on either intracellular β-glucosidase (GH1-1) or cellobiose phosphorylase (CBP), along with either an energy-consuming active cellodextrin transporter (CDT-1) or a non-energy-consuming passive cellodextrin facilitator (CDT-2). In this study, the ethanol production performance of two cellobiose-fermenting S. cerevisiae strains expressing mutant CDT-2 (N306I) with GH1-1 or CBP were compared with two cellobiose-fermenting S. cerevisiae strains expressing mutant CDT-1 (F213L) with GH1-1 or CBP in the simultaneous saccharification and fermentation (SSF) of cellulose under various conditions. It was found that, regardless of the SSF conditions, the phosphorolytic cellobiose-fermenting S. cerevisiae expressing mutant CDT-2 with CBP showed the best ethanol production among the four strains. In addition, during SSF contaminated by lactic acid bacteria (LAB), the phosphorolytic cellobiose-fermenting S. cerevisiae expressing mutant CDT-2 with CBP showed the highest ethanol production and the lowest lactate formation compared with those of other strains, such as the hydrolytic cellobiose-fermenting S. cerevisiae expressing mutant CDT-1 with GH1-1, and the glucose-fermenting S. cerevisiae with extracellular β-glucosidase. These results suggest that the cellobiose-fermenting yeast strain exhibiting low energy consumption can enhance the efficiency of the SSF of cellulosic biomass.

Keywords: Cellobiose phosphorylase; Cellulosic ethanol; Engineered Saccharomyces cerevisiae; Mutant cellodextrin facilitator; Simultaneous saccharification and fermentation.

Plants possess many glycoside hydrolase family 1 (GH1) β-glucosidases, which physiologically function in cell wall metabolism and activation of bioactive substances, but most remain uncharacterized. One GH1 isoenzyme AtBGlu42 in Arabidopsis thaliana has been identified to hydrolyze scopolin using the gene deficient plants, but no enzymatic properties were obtained. Its sequence similarity to another functionally characterized enzyme Os1BGlu4 in rice suggests that AtBGlu42 also acts on oligosaccharides. Here, we show that the recombinant AtBGlu42 possesses high kcat/Km not only on scopolin, but also on various β-glucosides, cellooligosaccharides, and laminarioligosaccharides. Of the cellooligosaccharides, cellotriose was the most preferred. The crystal structure, determined at 1.7 Å resolution, suggests that Arg342 gives unfavorable binding to cellooligosaccharides at subsite +3. The mutants R342Y and R342A showed the highest preference on cellotetraose or cellopentaose with increased affinities at subsite +3, indicating that the residues at this position have an important role for chain length specificity.

Keywords: Arabidopsis thaliana; X-ray crystallography; glycoside hydrolase family 1; substrate specificity; β-Glucosidase.

The San Francisco Bay Delta is experiencing seasonally warmer waters and salt water intrusion into historically freshwater ecosystems due to climate change. Steelhead/rainbow trout (Oncorhynchus mykiss) are resident in the Bay-Delta from juvenile development through the smoltification process. Due to sea level rise, premature seawater acclimation may co-occur with increased temperatures on pre-smolt juveniles. To evaluate the interactive effects of salinity and temperature on juvenile life stages of salmonids, rainbow trout alevin (3 days post-hatch) were exposed to 13^o C, 16.4^o C and 19^o C temperatures for 10 days and then challenged for 24 hours to 18 ppt seawater (SW). Similarly, fry (4 weeks post-hatch) were exposed to 13^o C, 16.4^o C and 19^o C temperatures for two weeks (14 days) and then challenged to SW. Estradiol-17β (E₂ ), cortisol, triiodothyronine (T₃ ), and thyroxine (T₄ ) were measured in whole animal homogenates and muscle tissue using Enzyme Linked Immunnosorbent Assays. Transcripts of gill Na⁺ /K⁺ ATPase β (NKAα1b), brain Growth Hormone I (gh1), and brain Gonadotropin-Releasing Hormone Receptor 2 (gnrh2) were also measured. Alevin exhibited a significant temperature-dependent decrease in survival and fry showed a temperature-dependent decrease in condition factor. Gene expression of NKAα1b, gh1, and gnrh2 was significantly decreased in all SW challenged alevin, and a significant decrease in gnrh2 expression was observed in fry with temperature. Alevin T₃ and T₄ concentrations were significantly increased with increasing temperature. There was a temperature-dependent increase in E₂ of fry, but not in alevin. The results of this study demonstrate that increasing temperature and SW exposure may adversely affect the survival and SW acclimation of alevin and fry stages of salmonids, and that the tolerances of younger juvenile stages should be considered when assessing the response of salmonid populations to climate change stressors. This article is protected by copyright. All rights reserved.

Keywords: Climate Change; Endocrine; Salmonids; Smoltification; Temperature; Thyroid Hormones; Trout.

This study aimed to investigate the effects of polymorphisms in growth hormone 1 (GH1) gene on the growth traits in Chinese indigenous yak. Using the polymerase chain reaction-restriction fragment length polymorphism (PCR-PFLP) approach, one novel single-nucleotide polymorphism (SNP), termed as g.1721G>A, was identified in the exon 4 of GH1 gene in 423 individuals of yak population. Based on the chi-square (χ²) test, the frequencies of g.1721G>A alleles agreed with Hardy-Weinberg equilibrium (HWE) (P < 0.05). A significant association was observed between this SNP and several growth traits (P < 0.01 or P < 0.05), in which the genotype GG exhibited the best values. The present study suggested that the identified SNP was a useful genetic marker for the improvement of growth traits in Chinese indigenous yak.

Keywords: Growth hormone 1 gene; Growth traits; Molecular characterization; Single-nucleotide polymorphism.

Bacteriocin-like inhibitory substances (BLIS) produced by Lactococcus lactis Gh1 had shown antimicrobial activity against Listeria monocytogenes ATCC 15313. Brain Heart Infusion (BHI) broth is used for the cultivation and enumeration of lactic acid bacteria, but there is a need to improve the current medium composition for enhancement of BLIS production, and one of the approaches is to model the optimization process and identify the most appropriate medium formulation. Response surface methodology (RSM) and artificial neural network (ANN) models were employed in this study. In medium optimization, ANN (R² = 0.98) methodology provided better estimation point and data fitting as compared to RSM (R² = 0.79). In ANN, the optimal medium consisted of 35.38 g/L soytone, 16 g/L fructose, 3.25 g/L sodium chloride (NaCl) and 5.40 g/L disodium phosphate (Na₂HPO₄). BLIS production in optimal medium (717.13 ± 0.76 AU/mL) was about 1.40-fold higher than that obtained in nonoptimised (520.56 ± 3.37 AU/mL) medium. BLIS production was further improved by about 1.18 times higher in 2 L stirred tank bioreactor (787.40 ± 1.30 AU/mL) as compared to that obtained in 250 mL shake flask (665.28 ± 14.22 AU/mL) using the optimised medium.

Keywords: Lactococcus lactis Gh1; artificial neural network; bacteriocin-like inhibitory substances; optimization; response surface methodology.

Background: There are many factors that lead to dwarfism, and the mechanism has not yet been elucidated. Next-generation sequencing may identify candidate-related gene mutations, which may clarify the molecular cause.

Aim: To analyze genetic variation by using a constructed panel related to dwarfism by utilizing next-generation sequencing platform sequencing analysis to screen candidate-related gene mutations.

Methods: Physical and laboratory characteristics, including clinical examination, growth hormone drug challenge test, serum insulin-like growth factor-1 (IGF-1), IGF binding protein 3, other related tests, imaging examination, and chromosome karyotyping, were analyzed. Next-generation sequencing was performed to analyze pathogenicity variability.

Results: In the 39 dwarfism patients, 10 had pathogenicity variability. Gene variation was found in the OBSL1, SLC26A2, PTPN11, COL27AI, HDAC6, CUL7, FGFR3, DYNC2H1, GH1, and ATP7B genes. Of the 10 patients with pathogenicity variability, the related physical characteristics included double breast development and growth hormone deficiency, enuresis and indirect inguinal hernia on the left, two finger distance of 70.2 cm, head circumference of 49.2 cm, ischium/lower body length of 1.8 cm, weak limb muscles, and partial growth hormone deficiency. After 6 mo of growth hormone therapy, the concentrations of IGF-1 and IGF binding protein 3 increased from 215.2 ± 170.3 to 285.0 ± 166.0 and 3.9 ± 1.4 to 4.2 ± 1.1, respectively.

Conclusion: OBSL1, SLC26A2, PTPN11, COL27AI, HDAC6, CUL7, FGFR3, DYNC2H1, GH1, and ATP7B genes may be related to the incidence of dwarfism, and more research needs to be performed to elucidate the mechanism.

Keywords: Dwarfism; Growth hormone; Mechanism; Next-generation sequencing; Pathogenicity variability; Therapy.

The role and mechanism of elagitannase is misunderstood because it exhibited different activities due to the low purity or complexity of substrates, and there is no available information about the biochemical, physicochemical and molecular characteristics of the enzyme. This study was aimed to obtain enzymatic extracts by Aspergillus niger GH1 in solid-state fermentation, using dextrose and ellagitannins as inducers of ellagitannase. Protein and bioinformatic analysis were performed to identify the protein sequence expressed in terms of culture conditions. The presence of ellagitannins increased ellagitannase activity 1143-fold compared to dextrose. The higher ellagitannase activity was found at 18 h of culture (1143.30 U g^-1PE). Three groups of proteins were identified in both cultures: β-glucosidase, phospholipase C, and triacylglycerol lipase. However, only phospholipase C was overexpressed with ellagitannins as inducers, showing the most spontaneous reaction with punicalagin (ΔG -8.56). These results suggest that phospholipase could be involved in ellagitannins biosynthesis.

Keywords: Ellagic acid; Ellagitannase; Protein analysis; Solid-state fermentation.

Two bacterial strains, designated SS33^T and Y03^T, were isolated from marine sediment and marine red alga collected on the coast of Weihai, PR China. Based on the results of 16S rRNA gene sequence analysis, strain SS33^T was found to be closely related to Primorskyibacter marinus PX7^T, Pelagivirga dicentrarchi YLY04^T, Palleronia marisminoris DSM 26347^T and Maribius pontilimi GH1-23^T with 94.8, 94.6, 94.5 and 94.5 % sequence similarity; strain Y03^T was found to be closest to Flavivirga aquimarina EC2D5^T, Flavivirga eckloniae ECD14^T and Flavivirga amylovorans JC2681^T with 96.4, 96.1 and 96.0 % sequence similarity. Strain SS33^T grew at 4-37 °C (optimum, 33 °C), at pH 6.0-9.5 (optimum, pH 7.5-8.0) and in the presence of 0-10 % (w/v) NaCl (optimum, 3.0 %). Chemotaxonomic analysis of strain SS33^T showed that the predominant respiratory quinone was ubiquinone-10. The major fatty acids (>10.0 %) included C_{18 : 1} ωc and C_{16 : 0}. The major polar lipids included phosphatidylglycerol, phosphatidylcholine, one unidentified phospholipid, one unidentified glycolipid, one unidentified polar lipid and two unidentified aminolipids. Strain Y03^T grew at 15-40 °C (optimum, 28 °C), at pH 6.5-8.0 (optimum, pH 7.0) and in the presence of 0.5-9.0 % (w/v) NaCl (optimum, 2.0%). Chemotaxonomic analysis showed that the predominant respiratory quinone was menaquinone-6. The major fatty acids (>10.0 %) included iso-C_{15 : 0}, iso-C_{15 : 1} G, iso-C_{17 : 0} 3-OH and iso-C_{15 : 0} 3-OH. The major polar lipids included phosphatidylethanolamine, one unidentified phospholipid, one unidentified aminolipid and four unidentified polar lipids. Based on the polyphasic data, strain SS33^T is considered to represent a novel species of the genus Palleronia, for which the name Palleronia sediminis sp. nov. is proposed, with the type strain SS33^T (=KCTC 62986^T=MCCC 1H00387^T). Strain Y03^T is considered to represent a novel species of the genus Flavivirga, for which the name Flavivirga algicola sp. nov. is proposed, with the type strain Y03^T (=KCTC 72001^T=MCCC 1H00386^T).

Keywords: 16S rRNA gene; Flavivirga algicola; Palleronia sediminis; Rhodobacteraceae; draft genome; polyphasic taxonomy.

Background: Isolated growth hormone deficiency (IGHD) due to mutations in GH1 gene is a rare disease caused by deficient production of endogenous growth hormone (GH).

Methods: We reported the clinical manifestation and genetic diagnosis (whole exome sequencing [WES], nested PCR Sanger sequencing, and rtPCR) of a family with two children with IGHD type I. We conducted a systematic review of cases with IGHD and compared height, and treatment outcomes in subtypes of IGHD.

Results: The patients were siblings born of nonconsanguineous parents from the Chinese Han population. The siblings both presented significantly short stature without other apparent abnormalities. The patients carry compound heterozygous mutations in GH1: a deletion and c.456 + 1G > A mutation that led to abnormal splicing. The systematic review identified 365 IGHD cases with GH1 mutations. Among these patients, their body height was most severely impaired in patients with IGHD type Ia, and the height standard deviation score decreased with the age of diagnosis in IGHD type Ia. Patients with IGHD type II had the longest duration of rhGH treatment, while patients with IGHD type Ib had the highest relative height improvement.

Conclusion: We identified two patients with IGHD type I caused by compound heterozygotic GH1 deletion and splicing mutation. The analysis of previously published IGHD patients suggests differences in linear growth among subtypes of IGHD.

Keywords: GH1; Growth hormone; IGHD; WES.

Glucose-tolerant and/or glucose-stimulated β-glucosidase is of great interest for its industrial utilization in enzymatic digestion of lignocellulosic biomass for biofuel production. In this study, a new gene of β-glucosidase MaGlu1A was cloned from an alginate-degrading marine bacterium Microbulbifer sp. ALW1. The gene of MaGlu1A encoded a 472-amino acid protein classified into the glycosyl hydrolase family 1 (GH1). The recombinant β-glucosidase was overexpressed and purified from Escherichia coli with a molecular mass of 65.0 kDa. Structure analysis illustrated the catalytic acid/base residue Glu186 and nucleophilic residue Glu370 in the enzyme. MaGlu1A displayed optimal activity at 40 °C and pH 4.5, respectively. It had substrate preference to the aryl-β-glycosidic bonds with glucose, fucose, and galactose moieties, in addition to cellobiose. MaGlu1A demonstrated strong stimulation to the supplemental glucose. Site-directed mutagenesis suggested an essential role of Asn242 in glucose stimulation. The enzymatic characterization of MaGlu1A provides general information about its catalytic properties facilitating its practical applications.

Keywords: Cellobiose digestion; Glucose stimulation; Microbulbifer sp; β-glucosidase.

Bacterial glycoside hydrolase 1 (GH1) enzymes with 6-phospho-β-galactosidase and 6-phospho-β-glucosidase activities have the important task of releasing phosphorylated and nonphosphorylated monosaccharides into the cytoplasm. Curiously, dual 6-phospho-β-galactosidase/6-phospho-β-glucosidase (dual-phospho) enzymes have broad specificity and are able to hydrolyze galacto- and gluco-derived substrates. This study investigates the structure and substrate specificity of a GH family 1 enzyme from Bacillus licheniformis, hereafter known as BlBglC. The enzyme structure has been solved, and sequence analysis, molecular dynamics simulations, and binding free energy calculations offered evidence of dual-phospho activity. Both test ligands p-nitrophenyl-β-d-galactoside-6-phosphate (PNP6Pgal) and p-nitrophenyl-β-d-glucoside-6-phosphate (PNP6Pglc) demonstrated strong binding to BlBglC although the pose and interactions of the PNP6Pglc triplicates were slightly more consistent. Interestingly, known specificity-inducing residues, Gln23 and Trp433, bind strongly to the ligand O3 hydroxyl group in the PNP6Pgal-BlBglC complex and to the ligand O4 hydroxyl group in the PNP6Pglc-BlBglC complex. Additionally, the BlBglC-His124 residue is a major contributor of hydrogen bonds to the PNP6Pgal O3 hydroxyl group but does not form any hydrogen bonds with PNP6Pglc. On the other hand, BlBglC residues Tyr173, Tyr301, Gln302, and Thr321 form hydrogen bonds with PNP6Pglc but not PNP6Pgal. These findings provide important details of the broad specificity of dual-phospho activity GH1 enzymes.

Background: A case of isolated growth hormone deficiency type IA (IGHD IA) caused by novel compound heterozygous mutation in the GH1 gene was reported in this study, which aimed to provide insights that will benefit future diagnosis and treatment.

Case presentation: We analyzed and summarized the clinical data and genetic test results from a patient with IGHD admitted in March 2019 to the Department of Pediatrics Tongji Hospital, Tongji Medical College, Huazhong University of Science and Technology. We described the results from a 1-year-9-months old female, whose chief complaint was "growth retardation for more than one year". Her birth length was 49.0 cm, and her birth weight was 3.05 kg. Suboptimal intake (breastfeeding) jaundice lasted for approximately two months following birth. When evaluated at the age of 1-year-9-months old, the patient's height was 61.0 cm (- 7.24 SD), and her weight was 6.4 kg (- 1.50 SD). The patient's physical characteristics included yellowish hair, large and unclosed anterior fontanelles, raised forehead, and a low and flat nose. The major abnormalities observed from the auxiliary examinations included low GH (< 0.05 μg/l), low IGF-1 (16.99 μg/l), and elevated TSH (6.97 mIU/l). Genetic testing revealed two heterozygous variants: a splicing mutation (NG_011676.1(NM_022560.4): c.10 + 1G>T, inherited from her mother) in intron 1 of the GH1 gene and a deletion that encompassed the same gene (chr17: 61973811-61996255, inherited from her father). After hormone replacement therapy with L-thyroxine and recombinant human GH (rhGH), the patient's thyroid function returned to normal, and her serum IGF-1 level significantly improved, which resulted in an accelerated increase in height.

Conclusion: This study described a case of IGHD caused by novel compound heterozygous mutations in the GH1 gene. This study suggested that closer attention should be directed to genetic testing and diagnosis based on clinical characteristics to avoid misdiagnosis.

Keywords: GH1; Gene; Isolated growth hormone deficiency; Mutation.

Context: Identification and biological actions of pituitary-derived exosomes remain elusive.

Objective: To validate production of exosomes derived from human and rat pituitary and elucidate their actions.

Methods: Isolated extracellular vesicles (EVs) were analyzed by Nanoparticle Tracking Analysis (NTA) and expressed exosomal markers detected by Western blot, using non-pituitary fibroblast FR and myoblast H9C2 cells as controls. Exosome inhibitor GW4869 was employed to detect attenuated EV release. Exosomal RNA contents were characterized by RNA-seq. In vitro and in vivo hepatocyte signaling alterations responding to GH1-derived exosomes (GH1-exo) were delineated by mRNA-seq. GH1-exo actions on protein synthesis, cAMP response, cell motility and metastases were assessed.

Results: NTA, exosomal marker detection, and GW4869 attenuated EV release, confirming the exosomal identity of pituitary EVs. Hydrocortisone increased exosome secretion in GH1 and GH3 cells, suggesting a stress-associated response. Exosomal RNA contents showed profiles distinct for pituitary cells, and rat primary hepatocytes exposed to GH1-exo exhibited transcriptomic alterations distinct from those elicited by GH or PRL. Intravenous GH1-exo injection into rats attenuated hepatic Eif2ak2 and Atf4 mRNA expression, both involved in cAMP responses and amino acid biosynthesis. GH1-exo suppressed protein synthesis and forskolin-induced cAMP levels in hepatocytes. GH1-exo treated HCT116 cells showed dysregulated p53 and MAPK pathways and attenuated motility of malignant HCT116 cells, and decreased tumor metastases in nude mice harboring splenic HCT116 implants.

Conclusions: Our findings elucidate biological actions of somatotroph-derived exosomes and implicate exosomes as non-hormonal pituitary-derived messengers.

Keywords: Exosome; nonhormonal actions; pituitary somatotroph adenoma.

We report four allelic variants (three novel) in three genes previously established as causal for hypopituitarism or related disorders. A novel homozygous variant in the growth hormone gene, GH1 c.171delT (p.Phe 57Leufs*43), was found in a male patient with severe isolated growth hormone deficiency (IGHD) born to consanguineous parents. A hemizygous SOX3 allelic variant (p.Met304Ile) was found in a male patient with IGHD and hypoplastic anterior pituitary. YASARA, a tool to evaluate protein stability, suggests that p.Met304Ile destabilizes the SOX3 protein (ΔΔG = 2.49 kcal/mol). A rare, heterozygous missense variant in the TALE homeobox protein gene, TGIF1 (c.268C>T:p.Arg90Cys) was found in a patient with combined pituitary hormone deficiency (CPHD), diabetes insipidus, and syndromic features of holoprosencephaly (HPE). This variant was previously reported in a patient with severe holoprosencephaly and shown to affect TGIF1 function. A novel heterozygous TGIF1 variant (c.82T>C:p.Ser28Pro) was identified in a patient with CPHD, pituitary aplasia and ectopic posterior lobe. Both TGIF1 variants have an autosomal dominant pattern of inheritance with incomplete penetrance. In conclusion, we have found allelic variants in three genes in hypopituitarism patients. We discuss these variants and associated patient phenotypes in relation to previously reported variants in these genes, expanding our knowledge of the phenotypic spectrum in patient populations.

Keywords: GH1; SOX3; TGIF1; allelic variants; hypopituitarism.

Alkyl glycosides are well-characterized nonionic surfactants, and can be prepared by transglycosylation reactions with retaining GH1 glycosidases being normally used for this purpose. The produced alkyl glycosides can also be hydrolyzed by the glycosidase, and hence, the yields of alkyl glycosides can be too low for industrial use. To improve the transglycosylation-to-hydrolysis ratio for a β-glucosidase from Thermotoga maritima (TmBglA) for the synthesis of alkyl glycoside, six mutants (N222F, N223C, N223Q, G224A, Y295F, and F414S) were produced. N222F, N223C, N223Q, G224A improved catalytic activity, F295Y and F414S are hydrolytically crippled with p-nitrophenol-β-d-glucopyranoside (pNPG) as substrate with an 85 and 70-fold decrease in apparent kcat, respectively; N222F shows the highest kcat/km value for pNPG. The substrate selectivity altered from pNPG to pNP-β-d-fucoside for N222F, F295Y, and F414S and from cellubiose to gentiobiose for N222F and F414S. Using pNPG (34 mM) and hexanol 80% (vol/vol), N222F, Y295F, and F414S synthesized hexyl-β-glycoside (HG) yields of 84.7%, 50.9%, and 54.1%, respectively, HG increased from 14.49 (TmBglA) to 22.8 mM (N222F) at 2 hr by 57.42%. However, this higher transglycosylation effect depended on that three mutants creates an environment more suited for hexanol in the active site pocket, and consequently suppressed its HG hydrolysis.

Keywords: Thermotoga maritima; Alkyl-glucosides; Transglycosylation mutants; β-Glucosidase.

<i>Aspergillus aculeatus</i> ZC-1005 (ZC-1005 was used as the abbreviation of this strain) is a hemicellulase-producing strain isolated from rotten citrus rind buried in the soil. Our previous study has shown its biochemical properties including high xylanase activity, mannanase activity, and degradation reaction with citrus mesocarp. In this study, we focused more on the enzyme safety evaluation and the genome sequencing <i>via</i> PacBio and Illumina platforms. High biological safety of the crude enzymes of ZC-1005 has been proven by the acute oral toxicity test, sub-chronic toxicity test, micronucleus test, and sperm malformation test. The genome of ZC-1005 had a GC content of 52.53%, with a size of 35,458,484 bp, and encoded 10,147 genes. Strain ZC-1005 harbored 269 glycosyl hydrolase (GH) genes of 64 families. The fungus produces cellulose-acting (GH3, GH5, <em>GH1</em>2, and <em>GH1</em>) and hemicellulose-acting enzymes (<em>GH1</em>6, GH31, GH2, and GH92). In genome annotation, we paid more attention to the genes encoding xylanase, such as gene 01512, gene 05833, gene 05469, gene 07781, gene 08432, gene 09042, gene 08008, and gene 09694. The collaboration between complete genome information and the degradation test confirmed that ZC-1005 could degrade cellulose and xylan. Our results showed that the citrus enzymatic decapsulation technology was efficacious and safe for canned citrus product processing, which may also solve the industrial waste problem. Therefore, ZC-1005 and the crude enzyme secreted from the strain were very promising to be used in the citrus processing industry.

Keywords: Aspergillus aculeatus ZC-1005; degradation; genome sequencing; safety evaluation; xylanase.

Gentiooligosaccharides (GnOS) are a kind of oligosaccharide formed by glucose with β-1-6 glycosidic bonds, which has become a new type of functional oligosaccharide for its unique refreshing bitter taste and valuable probiotic effects. However, the research on the enzymatic preparation of GnOS is not thorough enough. In this study, a GH1 thermophilic β-glucosidase from Thermotoga sp. KOL6 was used as a biocatalyst for the synthesis of GnOS. TsBgl1 exhibited excellent thermophilic and thermostable properties by possessing a melting temperature of 101.5 °C and reacting at 80-90 °C efficiently. Its half-life at 90 °C was approximately 5 h, suggesting its high heat resistance as well. TsBgl1 also showed excellent glucose tolerance with an inhibition constant (K_i) of 1720 mM and was stimulated in the presence of 0-900 mM glucose. TsBgl1 showed the highest hydrolytic activity on laminaribiose (Glc-β-1,3-Glc), but mainly synthetized gentiobiose (Glc-β-1,6-Glc) during transglycosylation. By optimizing the reaction conditions and substrate concentration, the highest yield of GnOS synthesized by TsBgl1 reached 144.3 g·L^-1 when 1000 g·L^-1 glucose was used as a substrate, which was higher than the highest yield ever reported. The thermophilic and thermostable properties of TsBgl1 were considered to be significant advantages in the industrial production of GnOS, where long periods of high-temperature reactions are required. This study was expected to provide an excellent candidate enzyme for industrial production of GnOS and also provide a reference for studying the transglycosylation of GH1 β-glucosidases.

Keywords: gentiobiose; gentiooligosaccharides; thermophilic; transglycosylation; β-glucosidase.

The study aims to elucidate the mechanisms responsible for the bioconversion of oleuropein into low molecular weight phenolic compounds in two selected L. plantarum strains, namely C11C8 and F3.5, under stress brine conditions and at two different temperatures (16°C and 30°C). For this purpose, we adopted an experimental strategy that combined high-resolution mass spectrometry, in silico functional analysis of glycosidic hydrolases GH1 encoding candidate genes and gene expression study. Oleuropein hydrolysis products and underlying enzymatic steps were identified, as well as a novel putative bgl gene, using seven strains belonging to the same species as controls, was detected. According to metabolomic analysis, a new intermediate compound (decarboxymethyl dialdehydic form of oleuropein aglycone) was revealed. In addition, the strain C11C8 showed a decrease in oleuropein content higher than F3.5 strain (30% vs 15%) at the temperature of 16°C. The highest increase in hydroxytyrosol was depicted by the strain C11C8 at the temperature of 30°C. PCR assay and sequencing analysis revealed that both strains possess bglH_1, bglH_2 and bglH3 genes. Furthermore, RT-PCR assay showed that bglH_3 is the only gene transcribed in all tested conditions, while bglH_2 is switched off in strain C11C8 grown under cold temperature and no transcription was detected for gene bglH_1. BglH_3 gene encodes a 6-phospho-beta-glucosidase, suggesting how phospho-β-glucosidase activity could belong to the overall metabolic strategy undertaken by L. plantarum to survive in an environment poor in free sugars, like table olive. Importance In the present study, a new candidate gene bglH_3 responsible for the β-glucosidase positive phenotype in L. plantarum was detected, providing the basis for future marker-assisted selection of L. plantarum starter strains with β-glucosidase positive phenotype. Furthermore, the ability of selected strains to hydrolyse oleuropein at low temperature is important for application as starter culture on an industrial-scale.