Identification of a d -Arabinose-5-Phosphate Isomerase in the Gram-Positive Clostridium tetani

Abstract

ABSTRACT d -Arabinose-5-phosphate (A5P) isomerases (APIs) catalyze the interconversion of d -ribulose-5-phosphate and d -arabinose-5-phosphate. Various Gram-negative bacteria, such as the uropathogenic Escherichia coli strain CFT073, contain multiple API paralogs (KdsD, GutQ, KpsF, and c3406) that have been assigned various cellular functions. The d -arabinose-5-phosphate formed by these enzymes seems to play important roles in the biosynthesis of lipopolysaccharide (LPS) and group 2 K-antigen capsules, as well as in the regulation of the cellular d -glucitol uptake and uropathogenic infectivity/virulence. The genome of a Gram-positive pathogenic bacterium, Clostridium tetani , contains a gene encoding a putative API, C. tetani API (CtAPI), even though C. tetani lacks both LPS and capsid biosynthetic genes. To better understand the physiological role of d -arabinose-5-phosphate in this Gram-positive organism, recombinant CtAPI was purified and characterized. CtAPI displays biochemical characteristics similar to those of APIs from Gram-negative organisms and complements the API deficiency of an E. coli API knockout strain. Thus, CtAPI represents the first d -arabinose-5-phosphate isomerase to be identified and characterized from a Gram-positive bacterium. IMPORTANCE The genome of Clostridium tetani , a pathogenic Gram-positive bacterium and the causative agent of tetanus, contains a gene (the CtAPI gene) that shares high sequence similarity with those of genes encoding d -arabinose-5-phosphate isomerases. APIs play an important role within Gram-negative bacteria in d -arabinose-5-phosphate production for lipopolysaccharide biosynthesis, capsule formation, and regulation of cellular d -glucitol uptake. The significance of our research is in identifying and characterizing CtAPI, the first Gram-positive API. Our findings show that CtAPI is specific to the interconversion of arabinose-5-phosphate and ribulose-5-phosphate while having no activity with the other sugars and sugar phosphates tested. We have speculated a regulatory role for this API in C. tetani , an organism that does not produce lipopolysaccharide.