Review
Modular evolution of the purine biosynthetic pathway

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Abstract

Structural studies, sequence alignments, and biochemistry have provided new insights into the evolution of the purine biosynthetic pathway. The importance of chemistry, the binding of ribose 5-phosphate (common to all purine biosynthetic intermediates), and transient protein–protein interactions in channeling of chemically unstable intermediates have all been examined in the past few years.

Introduction

The purine biosynthetic pathway is nearly ubiquitous and results in the conversion of phosphoribosyl pyrophosphate (PRPP) to inosine monophosphate (IMP). The seminal experiments in the 1950s and 1960s of Buchanan and his co-workers 1, 2, 3 established, in pigeon liver extracts, that 10 enzymatic activities are required for the interconversion of PRPP to IMP. In contrast, in most microorganisms, 12 enzymatic activities are involved in effecting the same interconversion (Figure 1). Structures of all of the enzymes in the bacterial pathway except PurL are now available, with seven of these structures from the same organism, Escherichia coli (PurF, PurD, PurN, PurM, PurK, PurT and PurE). The availability of this structural information in conjunction with the information from the genome sequencing projects has provided some provocative observations leading to insight into the factors that could govern the evolution of a biosynthetic pathway.

In general, biosynthetic pathways are built by patchwork. The proteins in pathways are composed of domains with defined catalytic activities that are fused to generate new activities. The purine pathway is no exception. In this review, we will present evidence for three different factors that should be considered in the evolution of the purine biosynthetic enzymes. First, we consider the importance of chemistry in this process. Of the twelve enzymes in procaryotes, six utilize ATP, two utilize glutamine, and two utilize N10-formyl tetrahydrofolate (THF). The question can be raised as to which of these proteins share structural homology. Second, in contrast with most biosynthetic pathways, all of the substrates in this pathway share a ribose 5-phosphate (R5P) moiety upon which the purine base is built sequentially (Figure 1). The question can be raised as to whether there is a common R5P-binding motif amongst the 12 enzymes. Finally, many of the intermediates in the purine biosynthetic pathway are chemically unstable. The question can be raised as to whether the surfaces of proteins provide a mechanism for direct transfer of these chemically unstable intermediates between successive enzymes in the pathway. In this review, each of these factors will be described in turn, focusing on recent information obtained from structural, sequence, and biochemical analyses.

The nomenclature of the enzymes and the intermediates in the purine pathway are cumbersome. To limit this complexity, we will call each protein by its gene name in E. coli, as indicated in Figure 1. In addition, each of the intermediates will be described using the acronyms in Figure 1. Initially we will focus on the importance of chemistry and the binding of R5P in the evolution of this pathway.

Section snippets

Chemistry and ribose 5-phosphate binding: importance in pathway evolution?

Four of the six ATP-requiring enzymes (PurD, PurK, PurT, and PurC) utilize ATP to phosphorylate a carboxylate, activating it for nucleophilic attack (Figure 2a). In the case of PurD, PurK and PurT, the nucleophiles are nucleotides, whereas in the case of PurC, the nucleophile is aspartate (Figure 2a). We have recently solved the structures of PurD and PurK from E. coli 4••, 5•. The high degree of sequence homology between PurK and PurT guarantees that all three of these enzymes are structurally

Protein surfaces: importance in pathway evolution?

The third factor that may be of importance in the evolution of a pathway involves the surfaces of the proteins that catalyze the formation of chemically unstable intermediates. The model is that the product–protein complex of one enzyme would present a surface recognized by the subsequent enzyme in the pathway. A transient interaction between the two proteins would result in the direct transfer (channeling) of the metabolite between the two proteins. In the purine pathway, we have postulated

Conclusions

The structural revolution, sequence information, and biochemical methods in vitro and in vivo have now allowed us unprecedented insight into the evolution of a biosynthetic pathway. The importance of chemistry, substrate binding, and the surfaces of proteins have been suggested by recent advances. Future studies should allow more rigorous analysis of the importance of these factors in the purine biosynthetic pathway and in other pathways.

Acknowledgements

Work on purine biosynthesis in the authors’ laboratories is supported by a National Institutes of Health grant to J Stubbe (GM32191). Research conducted at the Cornell High Energy Synchrotron Source (CHESS) is supported by the National Science Foundation under award DMR-9311772, and the Macromolecular Diffraction at CHESS (MacCHESS) facility, is supported by award RR-01646 from the National Institutes of Health. SE Ealick is indebted to the WM Keck Foundation and the Lucille P Markey Charitable

References and recommended reading

Papers of particular interest, published within the annual period of review, have been highlighted as:

  • • of special interest

  • •• of outstanding interest

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    Present address: Department of Chemistry, Washington University, One Brookings Drive, Campus Box 1134, St Louis, MO 63130, USA

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