Affiliation:
1. Department of Biochemistry and Molecular Genetics
2. Department of Microbiology
3. Department of Chemistry, University of Alabama at Birmingham, Birmingham, Alabama 35294
Abstract
ABSTRACT
Bacillus anthracis
spores, the etiological agents of anthrax, possess a loosely fitting outer layer called the exosporium that is composed of a basal layer and an external hairlike nap. The filaments of the nap are formed by trimers of the collagenlike glycoprotein BclA. Multiple pentasaccharide and trisaccharide side chains are O linked to BclA. The nonreducing terminal residue of the pentasaccharide side chain is the unusual sugar anthrose. A plausible biosynthetic pathway for anthrose biosynthesis has been proposed, and an
antABCD
operon encoding four putative anthrose biosynthetic enzymes has been identified. In this study, we genetically and biochemically characterized the activities of these enzymes. We also used mutant
B. anthracis
strains to determine the effects on BclA glycosylation of individually inactivating the genes of the anthrose operon. The inactivation of
antA
resulted in the appearance of BclA pentasaccharides containing anthrose analogs possessing shorter side chains linked to the amino group of the sugar. The inactivation of
antB
resulted in BclA being replaced with only trisaccharides, suggesting that the enzyme encoded by the gene is a dTDP-β-
l
-rhamnose α-1,3-
l
-rhamnosyl transferase that attaches the fourth residue of the pentasaccharide side chain. The inactivation of
antC
and
antD
resulted in the disappearance of BclA pentasaccharides and the appearance of a tetrasaccharide lacking anthrose. These phenotypes are entirely consistent with the proposed roles for the
antABCD
-encoded enzymes in anthrose biosynthesis. Purified AntA was then shown to exhibit β-methylcrotonyl-coenzyme A (CoA) hydratase activity, as we predicted. Similarly, we confirmed that purified AntC had aminotransferase activity and that purified AntD displayed
N
-acyltransferase activity.
Publisher
American Society for Microbiology
Subject
Molecular Biology,Microbiology
Cited by
32 articles.
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