Allosteric regulation accompanied by oligomeric state changes of Trypanosoma brucei GMP reductase through cystathionine-\u03b2-synthase domain

Akira Imamura,Bruno Kilunga Kubata,Takuya Otani,Manatsu Tamura,Shigenori Nishimura,Tomoka Kobayashi,Tetsuya Okada,Robert P Rambo,Hikaru Mase,Katsuaki Inoue,Takashi Inui,Susumu Uchiyama,Kentaro Ishii

doi:10.1038/s41467-020-15611-3

Abstract

Guanosine 5′-monophosphate reductase (GMPR) is involved in the purine salvage pathway and is conserved throughout evolution. Nonetheless, the GMPR of Trypanosoma brucei (TbGMPR) includes a unique structure known as the cystathionine-β-synthase (CBS) domain, though the role of this domain is not fully understood. Here, we show that guanine and adenine nucleotides exert positive and negative effects, respectively, on TbGMPR activity by binding allosterically to the CBS domain. The present structural analyses revealed that TbGMPR forms an octamer that shows a transition between relaxed and twisted conformations in the absence and presence of guanine nucleotides, respectively, whereas the TbGMPR octamer dissociates into two tetramers when ATP is available instead of guanine nucleotides. These findings demonstrate that the CBS domain plays a key role in the allosteric regulation of TbGMPR by facilitating the transition of its oligomeric state depending on ligand nucleotide availability.

Highlights

Guanosine 5′-monophosphate reductase (GMPR) is involved in the purine salvage pathway and is conserved throughout evolution
Enzymes harboring a CBS domain usually change their activities in the presence of purine nucleotides[10,11,12]; we sought to examine whether purine nucleotides modify the activity of T. brucei GMPR (TbGMPR)
Kinetic analysis demonstrated that the initial reaction velocity of TbGMPR without ligand nucleotides showed a sigmoidal curve when plotted against the concentrations of guanosine 5ʹmonophosphate (GMP), and the plots were well-fitted to the Hill equation (Fig. 1c, open circles)

Summary

Introduction

Guanosine 5′-monophosphate reductase (GMPR) is involved in the purine salvage pathway and is conserved throughout evolution. Our previous studies have demonstrated that a purine nucleotide analog, ribavirin 5ʹ-monophosphate, acts as an inhibitor for both T. brucei GMPR (TbGMPR) and IMPDH (TbIMPDH), and shows an anti-trypanosomal effect in culture when provided in the nucleoside form, ribavirin[5,6] Both GMPRs and IMPDHs have strong similarities in amino acid sequence, and their catalytic domains share the common structure of a (β/α)[8] barrel, referred to as a TIM barrel[7]; these two enzymes are still distinctive each other by the presence or absence of an additional domain[8]. Our findings suggest that the change in the oligomeric state of TbGMPR is responsible for allosteric regulation by nucleotide binding to the CBS domain

Methods

Results

Conclusion