Aquifex aeolicus 3-Deoxy-d-manno-2-Octulosonic Acid 8-Phosphate Synthase: A New Class of KDO 8-P Synthase?

Abstract

The relationship between 3-deoxy-D-manno-2-octulosonic acid 8-phosphate (KDO 8-P) synthase and 3-deoxy-D-arabino-2-heptulosonic acid 7-phosphate (DAH 7-P) synthase has not been adequately addressed in the literature. Based on recent reports of a metal requiring KDO 8-P synthase and the newly solved X-ray crystal structures of both Escherichia coli KDO 8-P synthase and DAH 7-P synthase, we begin to address the evolutionary kinship between these catalytically similar enzymes. Using a maximum likelihood-based grouping of 29 KDO 8-P synthase sequences, we demonstrate the existence of a new class of KDO 8-P synthase, the members of which we propose to require a metal cofactor for catalysis. Similarly, we hypothesize a class of DAH 7-P synthase that does not have the metal requirement of the heretofore model E. coli enzyme. Based on this information and a careful investigation of the reported X-ray crystal structures, we also propose that KDO 8-P synthase and DAH 7-P synthase are the product of a divergent evolutionary process from a common ancestor.

New Insights into the Evolutionary Links Relating to the 3-Deoxy-D-arabino-heptulosonate 7-Phosphate Synthase Subfamilies

“…This unique feedback regulatory mechanism observed with the aroA and aroQ fusion proteins prompted us to re-examine the results from previous phylogenetic studies (8,9) that divided the microbial DAHPS family into two subfamilies. ClustalW (17) analysis of the amino acid sequence found that AroAI ␤ DAHPSs, to which aroA(Q) Bs and aroA(Q) Pg were assigned, appear to consist of two types of enzymes (Table 3), one with a shorter sequence defining a single DAHPS domain (type I, ϳ270 residues) and one with a longer sequence that contains a single DAHPS domain plus an extra segment with divergent sequences at either the N or C terminus (type II, ϳ350 residues).…”

“…Previously, several biochemical properties (7,8) have been separately proposed as the possible criteria characterizing the individual subfamilies. Nevertheless, as discussed earlier, none of these suggestions have been demonstrated to be convincing.…”

“…Three different properties, metal requirement, allosteric effectors, and substrate specificity have been separately proposed as the possible criteria characterizing the individual subfamilies (7,8). The metal requirement hypothesis (8) suggests that the AroAI ␤ enzymes are non-metalloenzymes, whereas the AroAI ␣ enzymes are metalloenzymes; however, recent studies demonstrate that some AroAI ␤ DAHPSs are metalloenzymes.…”

“…DAHPSs exist in both microorganisms and plants (2)(3)(4)(5)(6)(7)(8)(9). Utilizing a maximum likelihood-based grouping analysis, Birck and Woodard (8) divided 25 randomly chosen microbial DAHPS protein sequences into two classes (defined as Class I and Class II).…”

“…The present of an absolute conserved metal binding sequence motif in all DAHPS, thus far identified, suggests that all DAHPSs are indeed metalloenzymes (6). Based on the results of DAHPS from the Bacillus subtilis derivative strain 168 (12), the allosteric effector hypothesis (8) proposes that the AroAI ␤ enzymes are inhibited by the branch intermediates chorismate/prephenate; whereas, the AroAI ␣ enzymes are inhibited by the terminal amino acid products Phe/Tyr/Trp. However, some AroAI ␣ enzymes have been shown to only be inhibited by their terminal amino acid products (6).…”