A new family of type III polyketide synthases in Mycobacterium tuberculosis.
The Mycobacterium tuberculosis genome has revealed a remarkable array of polyketide synthases (PKSs); however, no polyketide product has been isolated thus far. Most of the PKS genes have been implicated in the biosynthesis of complex lipids. We report here the characterization of two novel type III PKSs from M. tuberculosis that are involved in the biosynthesis of long-chain alpha-pyrones. Measurement of steady-state kinetic parameters demonstrated that the catalytic efficiency of PKS18 protein was severalfold higher for long-chain acyl-coenzyme A substrates as compared with the small-chain precursors. The specificity of PKS18 and PKS11 proteins toward long-chain aliphatic acyl-coenzyme A (C12 to C20) substrates is unprecedented in the chalcone synthase (CHS) family of condensing enzymes. Based on comparative modeling studies, we propose that these proteins might have evolved by fusing the catalytic machinery of CHS and beta-ketoacyl synthases, the two evolutionarily related members with conserved thiolase fold. The mechanistic and structural importance of several active site residues, as predicted by our structural model, was investigated by performing site-directed mutagenesis. The functional identification of diverse catalytic activity in mycobacterial type III PKSs provide a fascinating example of metabolite divergence in CHS-like proteins.
Saxena
Priti
P
National Institute of Immunology, Aruna Asaf Ali Marg, New Delhi 110 067, India.
Yadav
Gitanjali
G
Mohanty
Debasisa
D
Gokhale
Rajesh S
RS
D000214:Acyl Coenzyme A / Q000378:metabolism; D001665:Binding Sites; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D003460:Crystallization; D004591:Electrophoresis, Polyacrylamide Gel; D015723:Gene Library; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D007700:Kinetics; D008958:Models, Molecular; D015394:Molecular Structure; D009097:Multienzyme Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D009169:Mycobacterium tuberculosis / Q000201:enzymology* / Q000235:genetics; D010802:Phylogeny; D017385:Sequence Homology; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D000214:Acyl Coenzyme A; D007527:Isoenzymes; D009097:Multienzyme Complexes