Search

Substrate binding is typically one of the rate-limiting steps preceding enzyme catalytic action during homogeneous reactions. However, interlacial-based enzyme catalysis on insoluble crystalline substrates, like cellulose, has additional bottlenecks of individual biopolymer chain decrystallization from the substrate interlace followed by its processive depolymerization to soluble sugars. This additional decrystallization step has ramifications on the role of enzyme-substrate binding and its relationship to overall catalytic efficiency. We found that altering the crystalline structure of cellulose from its native allomorph l,~ to Ill, results in 40-50% lower binding partition coefficient for fungal cellulases. but surprisingly, it enhanced hydrolytic activity on the latter allomorph. We developed a comprehensive kinetic model for processive cellulases acting on insoluble substrates to explain this anomalous finding. Our model predicts that a reduction in the effective binding affinity to the substrate coupled with an increase in the decrystallization procession rate of individual cellulose chains from the substrate surface into the enzyme active site can reproduce our anomalous experimental findings. [ABSTRACT FROM AUTHOR]

As an important seedling source, monospores closely associate with yields in nori farming. However, the molecular mechanism underlying differences in monospore production for different strains remains unknown. Comparative transcriptome analysis was performed to examine gene expression differences between the spore abundant wild-type strain (WT) and spore deficient mutant (Y1) of Pyropia chauhanii. The WT strain that produces monospores in abundance exhibited more differentially expressed genes (DEGs) in both number and higher fold-changes than the Y1 strain incapable of producing monospores, indicating that the specific regulation of genes is involved in monospore production. Three lists of DEGs were obtained between the two strains using intersection and displayed in Venn diagram: one expressed only in WT strain, another expressed only in Y1 strain, and the third shared in both strains. DEGs annotated as homologous genes of Arabidopsis thaliana in these 3 lists were curated for online functional enrichment analysis on Metascape website. Gene regulatory networks of WT were functionally enriched in the processing, proteolysis, and transport of proteins, especially within the small GTPase protein family, which might be account for the monospore production ability, whereas Y1 were functionally enriched in the metabolism of essential substance and utilization of indispensable energy, which might be account for the rapid growth of blades. We found the differentially enriched gene regulatory networks between strains might be the intrinsic mechanisms of the different monospore production traits. These findings provide novel insights into the genes and regulatory networks associated with monospore production abilities, which are essential for developing accurate breeding technologies for optimal release of monospores and increase of total nori production. [ABSTRACT FROM AUTHOR]

Resistance of silicone resin to heat and irradiation was studied. The chemical structure of silicone resin before and after being heated and irradiated was analyzed by Fourier Transform Infrared Spectroscopy (FT-IR). The thermal stability of silicone resin was investigated by thermogravimetric analysis (TG) and differential thermogravimetry analysis (DTG), and its improvement was indicated by TG and DTG analyses of silicone resin after being irradiated by 60Co γ-ray under different fluence. The surface morphology, mechanical properties, corrosion resistance and chemical resistance of silicone resin clear paints before and after being heated and irradiated were studied by SEM, EIS et al. [ABSTRACT FROM AUTHOR]

Yesso scallop-pathogenic Vibrio splendidus strain JZ6 was found to have the highest virulence at 10°C, while its pathogenicity was significantly reduced with increased temperature and completely incapacitated at 28°C. In the present study, comparative transcriptome analyses of JZ6 and another nonpathogenic V. splendidus strain, TZ19, were conducted at two crucial culture temperatures (10°C and 28°C) in order to determine the possible mechanism of temperature regulation of virulence. Comparisons among four libraries, constructed from JZ6 and TZ19 cultured at 10°C and 28°C (designated JZ6_10, JZ6_28, TZ19_10, and TZ19_28), revealed that 241 genes were possibly related to the increased virulence of JZ6 at 10°C. There were 10 genes, including 2 encoding Flp pilus assembly proteins (FlhG and VS_2437), 6 encoding proteins of the "Vibrio cholerae pathogenic cycle" (ToxS, CqsA, CqsS, RpoS, HapR, and Vsm), and 2 encoding proteins in the Sec-dependent pathway (SecE and FtsY), that were significantly upregulated in JZ6_10 (P<0.05) compared to those in JZ6_28, TZ19_10, and TZ19_28, which were supposed to be responsible for adhesion, quorum sensing, virulence, and protein secretion of V. splendidus. When cultured at 10°C, JZ6 cells were larger and tended to aggregate more than those cultured at 28°C. The virulence factor (extracellular metalloprotease) was also found to be highly expressed in the extracellular product (ECP) of JZ6 at 10°C, and this ECP exhibited obvious cytotoxicity to oyster primary hemocytes, A549 cells, and L929 cells. These results indicated that low temperatures (10°C) could enhance adhesion, activate the quorum sensing systems, upregulate virulence factor synthesis and secretion, and, lastly, increase the pathogenicity of JZ6. [ABSTRACT FROM AUTHOR]

Background: Neural-endocrine-immune (NEI) system is a major modulation network among the nervous, endocrine and immune system and weights greatly in maintaining homeostasis of organisms during stress and infection. Some microRNAs are found interacting with NEI system (designated NeurimmiRs), addressing swift modulations on immune system. The oyster Crassostrea gigas, as an intertidal bivalve, has evolved a primary NEI system. However, the knowledge about NeurimmiRs in oysters remains largely unknown. Results: Six small RNA libraries from haemocytes of oysters stimulated with acetylcholine (ACh) and norepinephrine (NE) were sequenced to identify neurotransmitter-responsive miRNAs and survey their immunomodulation roles. A total of 331 miRNAs (132 identified in the present study plus 199 identified previously) were subjected to expression analysis, and twenty-one and sixteen of them were found ACh- or NE-responsive, respectively (FDR < 0.05). Meanwhile, 21 miRNAs exhibited different expression pattern after ACh or NE stimulation. Consequently, 355 genes were predicted as putative targets of these neurotransmitter-responsive miRNAs in oyster. Through gene onthology analysis, multiple genes involved in death, immune system process and response to stimulus were annotated to be modulated by NeurimmiRs. Besides, a significant decrease in haemocyte phagocytosis and late-apoptosis or necrosis rate was observed after ACh and NE stimulation (p < 0.05) while early-apoptosis rate remained unchanged. Conclusions: A comprehensive immune-related network involving PRRs, intracellular receptors, signaling transducers and immune effectors was proposed to be modulated by ACh- and NE-responsive NeurimmiRs, which would be indispensable for oyster haemocytes to respond against stress and infection. Characterization of the NeurimmiRs would be an essential step to understand the NEI system of invertebrate and the adaptation mechanism of oyster. [ABSTRACT FROM AUTHOR]

Conversion of lignocellulose to biofuels is partly inefficient due to the deleterious impact of cellulose crystallinity on enzymatic saccharification. We demonstrate how the synergistic activity of cellulases was enhanced by altering the hydrogen bond network within crystalline cellulose fibrils. We provide a molecular-scale explanation of these phenomena through molecular dynamics (MD) simulations and enzymatic assays. Ammonia transformed the naturally occurring crystalline allomorph Iβ to IIII, which led to a decrease in the number of cellulose intrasheet hydrogen bonds and an increase in the number of intersheet hydrogen bonds. This rearrangement of the hydrogen bond network within cellulose IIII, which increased the number of solvent-exposed glucan chain hydrogen bonds with water by ~50%, was accompanied by enhanced saccharification rates by up to 5-fold (closest to amorphous cellulose) and 60-70% lower maximum surface-bound cellulase capacity. The enhancement in apparent cellulase activity was attributed to the "amorphous-like" nature of the cellulose IIII fibril surface that facilitated easier glucan chain extraction. Unrestricted substrate accessibility to active-site clefts of certain endocellulase families further accelerated deconstruction of cellulose IIII. Structural and dynamical features of cellulose IIII, revealed by MD simulations, gave additional insights into the role of cellulose crystal structure on fibril surface hydration that influences interfacial enzyme binding. Subtle alterations within the cellulose hydrogen bond network provide an attractive way to enhance its deconstruction and offer unique insight into the nature of cellulose recalcitrance. This approach can lead to unconventional pathways for development of novel pretreatments and engineered cellulases for cost-effective biofuels production. [ABSTRACT FROM AUTHOR]

The sfr6-1 mutant of Arabidopsis thaliana was identified previously on the basis of its failure to undergo acclimation to freezing temperatures following exposure to low positive temperatures. This failure is attributed to a defect in the pathway leading to cold on-regulated ( COR) gene expression via CBF (C-box binding factor) transcription factors. We identified a region of chromosome 4 containing SFR6 by positional mapping. Fine mapping of the sfr6-1 mutation proved impossible as the locus resides very close to the centromere. Therefore, we screened 380 T-DNA lines with insertions in genes within the large region to which sfr6-1 mapped. This resulted in the identification of two further mutant alleles of SFR6 ( sfr6-2 and sfr6-3); like the original sfr6-1 mutation, these disrupt freezing tolerance and COR gene expression. To determine the protein sequence, we cloned an SFR6 cDNA based on the predicted coding sequence, but this offered no indication as to the mechanism by which SFR6 acts. The SFR6 gene itself is not strongly regulated by cold, thus discounting regulation of SFR6 activity at the transcriptional level. We show that over-expression of CBF1 or CBF2 transcription factors, which constitutively activate COR genes in the wild-type, cannot do so in sfr6-1. We demonstrate that CBF protein accumulates to wild-type levels in response to cold in sfr6-1. These results indicate a role for the SFR6 protein in the CBF pathway -downstream of CBF translation. The fact that the SFR6 protein is targeted to the nucleus may suggest a direct role in modulating gene expression. [ABSTRACT FROM AUTHOR]