Formation of catalytically active S ‐acetyl(malonate decarboxylase) requires malonyl‐coenzyme A Acyl carrier protein transacylase as auxiliary enzyme

• Published in: European journal of biochemistry Vol. 259; no. 1-2; pp. 181 - 187
• Main Authors: Hoenke, Stefan, Dimroth, Peter
• Format: Journal Article
• Language: English
• Published: 01.01.1999
• ISSN: 0014-2956; 1432-1033
• DOI: 10.1046/j.1432-1327.1999.00034.x

Functional malonate decarboxylase of Klebsiella pneumoniae is an acetyl‐S‐enzyme with an acetylated phosphoribosyl dephospho‐CoA prosthetic group. The mdcH gene product acts as a malonyl‐CoA:ACP transacylase and initiates the activation of (deacetyl)malonate decarboxylase by malonyl‐transfer to the prosthetic group. The malonyl residue is subsequently decarboxylated to an acetyl residue by the decarboxylase itself. Purified malonate decarboxylase consists of the four subunits MdcA, D, E and C in an apparent 1 : 1 : 1 : 1 stoichiometry. In addition, the preparation contains substoichiometric amounts of MdcH comigrating on SDS/PAGE with MdcD. Malonate decarboxylase isolated from strains with a deletion of the mdcH gene was not activated with malonyl‐CoA. Activity could be gained, however, in the additional presence of MdcH that has been synthesized in Escherichia coli and purified from inclusion bodies. Substrates for MdcH are malonyl‐CoA or methylmalonyl‐CoA but not acetyl‐CoA. The enzyme has K m values of 16 µ m for both substrates and V max for malonyl‐CoA of 190 U·mg –1 and for methylmalonyl‐CoA of 37 U·mg –1 . Transfer of the methylmalonyl‐residue to the prosthetic group proceeds via the covalent methylmalonyl‐MdcH intermediate. The transacylase is specifically inhibited by N ‐ethylmaleimide, and preincubation with malonyl‐CoA or methylmalonyl‐CoA protects the enzyme from this inhibition.