β‐Glucosidase, β‐galactosidase, family A cellulases, family F xylanases and two barley glycanases form a superfamily of enzymes wit 8‐fold β/α architecture and with two conserved glutamates near the carboxy‐terminal ends of β‐strands four and seven

Jenkins, John A.; Leggio, Leila Lo; Harris, Gillian W.; Pickersgill, Richard W.

doi:10.1016/0014-5793(95)00252-5

Cited by 256 publications

(186 citation statements)

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“…The members of the family also belong to the 4/7-superfamily. An important characteristic of the superfamily is the presence of aspartates/glutamates at the C terminal of the fourth and seventh strands of the @/a)8-bamel (Jenkins et al, 1995;Wiesmann et al, 1995). The relevant residues in GTF Asp 415 and Asp 526 are located close to the C terminus of the P-strand four and P-strand seven.…”

Section: Site-directed Mutagenesis Of Putative Gtf Catalytic Residuesmentioning

confidence: 99%

Knowledge‐based model of a glucosyltransferase from the oral bacterial group of mutans streptococci

et al. 1997

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Mutans streptococci glucosyltransferases catalyze glucosyl transfer from sucrose to a glucan chain. We previously identified an aspartyl residue that participates in stabilizing the glucosyl transition state. The sequence surrounding the aspartate was found to have substantial sequence similarity with members of a-amylase family. Because little is known of the protein structure beyond the amino acid sequence, we used a knowledge-based interactive algorithm, MACAW, which provided significant level of homology with a-amylases and glucosyltransferase from Streptococcus downei gtfl (GTF). The significance of GTF similarity is underlined by GTF/a-amylase residues conserved in all but one a-amylase invariant residues. Site-directed mutagenesis of the three GTF catalytic residues are homologous with the a-amylase catalytic triad. The glucosyltransferases are members of the 4/7-superfamily that have a (P/a)8-barrel structure and belong to family 13 of the glycohydralases.Keywords: (P/a)x-barrel; catalytic residues; enzymes; mutans streptococci glucosyltransferase; protein modeling; site-directed mutagenesis Oral bacteria referred to as mutans streptococci secrete glucosyltransferases (EC2.4.1.5), which are a significant virulence factor in initiation of dental caries on smooth enamel surfaces and cementa1 root surfaces (Hamada & Slade, 1980;Loesche, 1986). The extracellular enzymes catalyze glucosyl transfer from sucrose to a growing glucan chain. The bacterial colony produces metabolic acids that demineralize the tooth surface and, if unchecked, will lead to dental caries.GTFs are very large proteins of approximately 1,300-1,700 residues. The enzyme can be grossly divided into an N-terminal catalytic domain and a C-terminal glucan-binding domain that captures the synthesized polysaccharide (Ferretti et al., Wong et al., 1990). The large size of the enzyme has made it difficult to develop structure information beyond the solution of the amino acid sequence. Nonetheless, we became aware of sequence similarity between GTF and members of the a-amylase family. In our earlier studies, we trapped a glucosyl-enzyme catalytic intermediate (Mooser & Iwaoka, 1989) that was coordinated with an aspartyl Reprint requests to: Gregory Mooser, Department of Basic Sciences, School of Dentistry, University of Southern California, Los Angeles, California 90089-0641; e-mail: gmooser@hsc.usc.edu.Abbreviations: GTF, glucosyltransferase from Streptococcus downei g@; MACAW, multiple alignment construction and analysis workbench; SP, sum of the pairs; dNJ, I-deoxynojirimycin. residue, and the sequence surrounding the aspartate was homologous with a sequence conserved in all members of the a-amylase family and several related enzymes (Mooser et al., Mooser, 1992).In this study, we coupled knowledge-based multiple sequence alignment with site-directed mutagenesis to establish evidence of structural similarity between GTF and solved crystal structures of members of the a-amylase family. The members of the GTF family are highly similar and ...

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Section: Site-directed Mutagenesis Of Putative Gtf Catalytic Residuesmentioning

confidence: 99%

Knowledge‐based model of a glucosyltransferase from the oral bacterial group of mutans streptococci

et al. 1997

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“…Some structural folds have given rise to a myriad of enzymes that display significant differences in specificity, exemplified by the GHs located in clan GH-A. Members of this clan display a (b/a) 8 -fold in which the catalytic residues are presented at the C terminus of b-strands 4 and 7 Jenkins et al, 1995). While the enzymes all hydrolyze an equatorial glycosidic bond, their mode of action (exo and endo), specificity for the sugar at the catalytic 21 subsite and more distal regions of the substratebinding region (Xyl, Man, Glc, Araf, Gal), and the linkage cleaved (e.g.…”

Section: Catalytic Modules Of Glycoside Hydrolasesmentioning

confidence: 99%

The Biochemistry and Structural Biology of Plant Cell Wall Deconstruction

2010

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“…The overall fold of the CBG molecule is a (b/a) 8 barrel. Based on the three-dimensional structure and sequence analysis of family 1, plant b-glucosidases also belong to the 4/7 superfamily [8]. The 4/7 superfamily consists of glycosyl hydrolases belonging to a superfamily of eightfold b/a-barrels with a nucleophile (usually E) located close to the C-terminus of b-strand 7, and a proton donor (usually E) close to the C-terminus of b-strand 4.…”

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confidence: 99%

“…The threedimensional structure of a cyanogenic b-glucosidase from white clover (CBG) has been elucidated recently [7]. The overall fold of the CBG molecule is a (b/a) 8 barrel. Based on the three-dimensional structure and sequence analysis of family 1, plant b-glucosidases also belong to the 4/7 superfamily [8].…”

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confidence: 99%

The role of cysteine residues in structure and enzyme activity of a maize β‐glucosidase

Rotrekl¹,

Nejedlá²,

Kučera³

et al. 1999

European Journal of Biochemistry

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The maize Zm-p60.1 gene encodes a b-glucosidase that can release active cytokinins from their storage forms, cytokinin-O-glucosides. Mature catalytically active Zm-p60.1 is a homodimer containing five cysteine residues per a subunit. Their role was studied by mutating them to alanine (A), serine (S), arginine (R) or aspartic acid (D) using site-directed mutagenesis, and subsequent heterologous expression in Escherichia coli. All substitutions of C205 and C211 resulted in decreased formation and/or stability of the homodimer, manifested as accumulation of high levels of monomer in the bacterial expression system. Examination of urea-and glutathione-induced dissociation patterns of the homodimer to the monomers, HPLC profiles of hydrolytic fragments of reduced and oxidized forms, and a homology-based three-dimensional structural model revealed that an intramolecular disulfide bridge formed between C205 and C211 within the subunits stabilized the quaternary structure of the enzyme. Mutating C52 to R produced a monomeric enzyme protein, too. No detectable effects on homodimer formation were apparent in C170 and C479 mutants. Given the K m values for C170A/S mutants were equal to that for the wild-type enzyme, C170 cannot participate in enzyme±substrate interactions. Possible indirect effects of C170A/S mutations on catalytic activity of the enzyme were inferred from slight decreases in the apparent catalytic activity, k H cat . C170 is located on a hydrophobic side of an a-helix packed against hydrophobic aminoacid residues of b-strand 4, indicating participation of C170 in stabilization of a (b/a) 8 barrel structure in the enzyme. In C479A/D/R/S mutants, K m and k H cat were influenced more significantly suggesting a role for C479 in enzyme catalytic action.Keywords: b-glucosidase; cysteine residues; disulfide bridge; structure±fuction relationships.Current research on b-glucosidases of animals, plants and microorganisms has significant scientific, clinical and economic implications (reviewed in [1]). In plants, b-glucosidases have been implicated in various aspects of growth, productivity and defence, and reactions such as cyanogenesis related to food and feed toxicity. In addition, recent data indicate that plant b-glucosidases may be involved in the metabolism of plant hormones, whose storage forms occur as b-glucosides and are activated by b-glucosidases [2,3]. Thus, information on structure±function relationships in the b-glucosidases is of obvious importance for understanding their biological functions, and engineering their catalytic and molecular properties for industrial and field applications.b-Glucosidases catalyse the hydrolysis of aryl and alkyl-b-dglucosides as well as glucosides with only carbohydrate moieties (such as cellobiose). Catalysis involves two essential carboxylates, one acting as a proton donor and the other as a nucleophile [4]. Cleavage of the glucosidic bond results in retention of the configuration at the anomeric carbon atom.Plant b-glucosidases are classified into family 1 of glycos...

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β‐Glucosidase, β‐galactosidase, family A cellulases, family F xylanases and two barley glycanases form a superfamily of enzymes wit 8‐fold β/α architecture and with two conserved glutamates near the carboxy‐terminal ends of β‐strands four and seven

Cited by 256 publications

References 36 publications

Knowledge‐based model of a glucosyltransferase from the oral bacterial group of mutans streptococci

Knowledge‐based model of a glucosyltransferase from the oral bacterial group of mutans streptococci

The Biochemistry and Structural Biology of Plant Cell Wall Deconstruction

The role of cysteine residues in structure and enzyme activity of a maize β‐glucosidase

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