Mandelate Pathway of Pseudomonas putida: Sequence Relationships Involving Mandelate Racemase, (S)-Mandelate Dehydrogenase, and Benzoylformate Decarboxylase and Expression of Benzoylformate Decarboxylase in Escherichia coli

Amy Y. Tsou, Stephen C. Ransom, John A. Gerlt, Douglas D. Buechter, Patricia C. Babbitt, George L. Kenyon

Research output: Contribution to journalArticle

Abstract

The genes that encode the five known enzymes of the mandelate pathway of Pseudomonas putida (ATCC 12633), mandelate racemase (mdlA), (S)-mandelate dehydrogenase (mdlB), benzoylformate decarboxylase (mdlC), NAD+-dependent benzaldehyde dehydrogenase (mdlD), and NADP+-dependent benzaldehyde dehydrogenase (mdlE), have been cloned. The genes for (S)-mandelate dehydrogenase and benzoylformate decarboxylase have been sequenced; these genes and that for mandelate racemase [Ransom, S. C., Gerlt, J. A., Powers, V. M., & Kenyon, G. L. (1988) Biochemistry 27, 540] are organized in an operon (mdlCBA). Mandelate racemase has regions of sequence similarity to muconate lactonizing enzymes I and II from P. putida. (S)-Mandelate dehydrogenase is predicted to be 393 amino acids in length and to have a molecular weight of 43 352; it has regions of sequence similarity to glycolate oxidase from spinach and ferricytochrome b2 lactate dehydrogenase from yeast. Benzoylformate decarboxylase is predicted to be 499 amino acids in length and to have a molecular weight of 53 621; it has regions of sequence similarity to enzymes that decarboxylate pyruvate with thiamin pyrophosphate as cofactor. These observations support the hypothesis that the mandelate pathway evolved by recruitment of enzymes from preexisting metabolic pathways. The gene for benzoylformate decarboxylase has been expressed in Escherichia coli with the trc promoter, and homogeneous enzyme has been isolated from induced cells.

Original languageEnglish (US)
Pages (from-to)9856-9862
Number of pages7
JournalBiochemistry
Volume29
Issue number42
DOIs
StatePublished - Oct 1 1990

ASJC Scopus subject areas

  • Biochemistry

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