Structural characterization of a ribose-5-phosphate isomerase B from the pathogenic fungus Coccidioides immitis

Thomas E Edwards,Bart L Staker,Anna S Gardberg,Jess T Leonard,David J Leibly,Wesley C Van Voorhis,Kaitlin B Thompkins,Ariel B Abramov,Lance J Stewart,Matthew C Clifton,Eric R Smith,Peter J Myler,Ruth O Baydo

doi:10.1186/1472-6807-11-39

Abstract

BackgroundRibose-5-phosphate isomerase is an enzyme that catalyzes the interconversion of ribose-5-phosphate and ribulose-5-phosphate. This family of enzymes naturally occurs in two distinct classes, RpiA and RpiB, which play an important role in the pentose phosphate pathway and nucleotide and co-factor biogenesis.ResultsAlthough RpiB occurs predominantly in bacteria, here we report crystal structures of a putative RpiB from the pathogenic fungus Coccidioides immitis. A 1.9 Å resolution apo structure was solved by combined molecular replacement and single wavelength anomalous dispersion (SAD) phasing using a crystal soaked briefly in a solution containing a high concentration of iodide ions. RpiB from C. immitis contains modest sequence and high structural homology to other known RpiB structures. A 1.8 Å resolution phosphate-bound structure demonstrates phosphate recognition and charge stabilization by a single positively charged residue whereas other members of this family use up to five positively charged residues to contact the phosphate of ribose-5-phosphate. A 1.7 Å resolution structure was obtained in which the catalytic base of C. immitis RpiB, Cys76, appears to form a weakly covalent bond with the central carbon of malonic acid with a bond distance of 2.2 Å. This interaction may mimic that formed by the suicide inhibitor iodoacetic acid with RpiB.ConclusionThe C. immitis RpiB contains the same fold and similar features as other members of this class of enzymes such as a highly reactive active site cysteine residue, but utilizes a divergent phosphate recognition strategy and may recognize a different substrate altogether.

Highlights

Ribose-5-phosphate isomerase is an enzyme that catalyzes the interconversion of ribose-5-phosphate and ribulose-5-phosphate
Structure determination of C. immitis RpiB Given that the C. immitis RpiB contains modest sequence identity to other structurally characterized RpiBs, we attempted to solve the crystal structure of C. immitis RpiB by iodide ion single wavelength anomalous dispersion (SAD) phasing [10,11], a strategy that has proven successful for structure determination of many SSGCID targets [12,13]
The partial molecular replacement solution was combined with the SAD phases and yielded a clearly interpretable electron density map (FOM 0.53 prior to density modification) into which Buccaneer [16] built both molecules in the asymmetric unit end-to-end in less than one minute (Table 1; Figure 1)

Summary

Introduction

Ribose-5-phosphate isomerase is an enzyme that catalyzes the interconversion of ribose-5-phosphate and ribulose-5-phosphate This family of enzymes naturally occurs in two distinct classes, RpiA and RpiB, which play an important role in the pentose phosphate pathway and nucleotide and co-factor biogenesis. Ribose-5-phosphate isomerases catalyze the interconversion of ribulose-5-phosphate and ribose-5-phosphate as an important part of the pentose phosphate pathway [1]. Two types of ribose-5-phosphate isomerases exist, RpiA and RpiB, which share little structural homology and have distinct active sites and mechanisms of action [2]. Most organisms such as Escherichia coli contain both RpiA and RpiB, but other organisms contain only one class of.

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Results

Conclusion