Biology subjects

Yamada, C.

Publications and source records attributed to Yamada, C..

3 recordsLinked to original sources

bioRxiv2020

Crystallographic and cryogenic electron microscopic structures and enzymatic characterization of sulfur oxygenase reductase from Sulfurisphaera tokodaii

Sulfur oxygenase reductases (SORs) are present in thermophilic and mesophilic archaea and bacteria, and catalyze oxygen-dependent oxygenation and disproportionation of elemental sulfur. SOR has a hollow, spherical homo-24-mer structure and reactions take place at active sites inside the chamber. The crystal structures of SORs from two Acidianus species have been reported. However, the states of the active site components (mononuclear iron and cysteines) and the entry and exit paths of the substrate and products are still in dispute. Here, we report the biochemical and structural characterizations of SORs from the thermoacidophilic archaeon Sulfurisphaera tokodaii (StSOR) and present high-resolution structures determined by X-ray crystallography and cryogenic electron microscopy (cryo-EM). The crystal structure of StSOR was determined at 1.73 [A] resolution. At the catalytic center, iron is ligated to His86, His90, Glu114, and two water molecules. Three conserved cysteines in the cavity are located 9.5[~]13 [A] from the iron and were observed as free thiol forms. A mutational analysis indicated that the iron and one of the cysteines (Cys31) were essential for both activities and the other two cysteines (Cys101 and Cys104) had a supportive role. The cryo-EM structure was determined at 2.24 [A] resolution using an instrument operating at 200 kV. The two structures determined by different methodologies showed similar main chain traces, but the maps exhibited different features at catalytically important components. Given the high resolution achieved in this study, StSOR was shown to be a good benchmark sample for cryo-EM measurements. HighlightsO_LISulfur oxygenase reductase (SOR) was biochemically and structurally characterized. C_LIO_LIHigh resolution structures of SOR were determined by crystallography and cryo-EM. C_LIO_LITwenty-four identical subunits of SOR form a hollow sphere. C_LIO_LICatalytic components exhibited different features in the crystal and cryo-EM structures. C_LI

biochemistry↗

bioRxiv2020

Butyrate producing Clostridiales utilize distinct human milk oligosaccharides correlating to early colonization and prevalence in the human gut

The early life human gut microbiota exerts life-long health effects on the host, but the mechanisms underpinning its assembly remain elusive. Particularly, the early colonization of Clostridiales from the Roseburia-Eubacterium group, associated with protection from colorectal cancer, immune- and metabolic disorders is enigmatic. Here we unveil the growth of Roseburia and Eubacterium members on human milk oligosaccharides (HMOs) using an unprecedented catabolic apparatus. The described HMO pathways and additional glycan utilization loci confer co-growth with Akkermansia muciniphilia via cross-feeding and access to mucin O-glycans. Strikingly, both, HMO and xylooligosaccharide pathways, were active simultaneously attesting an adaptation to a mixed HMO-solid food diet. Analyses of 4599 Roseburia genomes underscored the preponderance of HMO pathways and highlighted different HMO utilization phylotypes. Our revelations provide a possible rationale for the early establishment and resilience of butyrate producing Clostridiales and expand the role of specific HMOs in the assembly of the early life microbiota.

microbiology↗

bioRxiv2020

Crystal structure of β-L-arabinobiosidase belonging to glycoside hydrolase family 121

Enzymes acting on -L-arabinofuranosides have been extensively studied; however, the structures and functions of {beta}-L-arabinofuranosidases are not fully understood. Three enzymes and an ABC transporter in a gene cluster of Bifidobacterium longum JCM 1217 constitute a degradation and import system of {beta}-L-arabinooligosaccharides on plant hydroxyproline-rich glycoproteins. An extracellular {beta}-L-arabinobiosidase (HypBA2) belonging to the glycoside hydrolase (GH) family 121 plays a key role in the degradation pathway by releasing {beta}-1,2-linked arabinofuranose disaccharide ({beta}-Ara2) for the specific sugar importer. Here, we present the crystal structure of the catalytic region of HypBA2 as the first three-dimensional structure of GH121 at 1.85 [A] resolution. The HypBA2 structure consists of a central catalytic (/)6 barrel domain and two flanking (N- and C-terminal) {beta}-sandwich domains. A pocket in the catalytic domain appears to be suitable for accommodating the {beta}-Ara2 disaccharide; this pocket is highly conserved among GH121 proteins. The three acidic residues Glu383, Asp515, and Glu713, located in this pocket, are completely conserved among all ~270 members of GH121; site-directed mutagenesis analysis showed that they are essential for catalytic activity. The active site of HypBA2 was compared with those of GH63 -glycosidase, GH94 chitobiose phosphorylase, GH142 {beta}-L-arabinofuranosidase, GH78 -L-rhamnosidase, and GH37 ,-trehalase. Based on these analyses, we concluded that the three conserved residues are essential for catalysis and substrate binding. {beta}-L-Arabinobiosidase genes in GH121 are mainly found in the genomes of bifidobacteria and Xanthomonas species, suggesting that the cleavage and specific import system for the {beta}-Ara2 disaccharide on plant hydroxyproline-rich glycoproteins are shared in animal gut symbionts and plant pathogens.

biochemistry↗