Functional and Structural Analyses of a Highly Multifunctional Enzyme TM1270 from the Hyperthermophile Thermotoga maritima

Abstract

The hyperthermophile Thermotoga maritima possesses d-amino acid-metabolizing enzymes and multifunctional enzymes associated with l- and d-amino acid metabolism, although it does not have typical alanine and glutamate racemases. Intriguingly, in this study, we found that unexpectedly one PLP fold-type I enzyme from this organism, TM1270, has six different enzyme activities, namely amino acid racemase, cystathionine β-lyase, serine dehydratase, threonine aldolase, aspartate 4-decarboxylase, and amino acid aminotransferase activities. We characterized the properties of these six enzyme activities including their substrate specificities, pH and temperature dependences, and kinetic parameters. β-Lyase activity was the highest among the six activities based on kinetic parameters. Furthermore, we determined the crystal structure of TM1270 with the internal aldimine form of pyridoxal 5′-phosphate, which forms a Schiff base with Lys202. The possible reaction mechanisms of the six enzyme activities are proposed based on the crystal structure and the results of mutational analysis.