We investigate the evolution of co-occurring analogous enzymes involved in L-tryptophan and L-histidine biosynthesis in Actinobacteria. Phylogenetic analysis of trpF homologues, a missing gene in certain clades of this lineage whose absence is complemented by a dual-substrate HisA homologue, termed PriA, found that they fall into three categories: (i) trpF-1, an L-tryptophan biosynthetic gene horizontally acquired by certain Corynebacterium species; (ii) trpF-2, a paralogue known to be involved in synthesizing a pyrrolopyrrole moiety and (iii) trpF-3, a variable non-conserved orthologue of trpF-1. We previously investigated the effect of trpF-1 upon the evolution of PriA substrate specificity, but nothing is known about the relationship between trpF-3 and priA. After in vitro steady-state enzyme kinetics we found that trpF-3 encodes a phosphoribosyl anthranilate isomerase. However, mutation of this gene in Streptomyces sviceus did not lead to auxothrophy, as expected from the biosynthetic role of trpF-1. Biochemical characterization of a dozen co-occurring TrpF-2 or TrpF-3, with PriA homologues, explained the prototrophic phenotype, and unveiled an enzyme activity trade-off between TrpF and PriA. X-ray structural analysis suggests that the function of these PriA homologues is mediated by non-conserved mutations in the flexible L5 loop, which may be responsible for different substrate affinities. Thus, the PriA homologues that co-occur with TrpF-3 represent a novel enzyme family, termed PriB, which evolved in response to PRA isomerase activity. The characterization of co-occurring enzymes provides insights into the influence of functional redundancy on the evolution of enzyme function, which could be useful for enzyme functional annotation.
Bibliographical noteFunding Information:
This work was supported by the Conacyt Mexico [grant number 179290 (to F.B.-G.)]; the National Institutes of Health [grant number GM094585 (to A.J.)]; the U.S. Department of Energy, Office of Biological and Environmental Research [grant number DE-AC02-06CH11357].
© 2016 Authors; published by Portland Press Limited.
- (βα) -barrel
- Analogous enzymes
- Substrate specificity