Prediction and manipulation of the stereochemistry of enoylreduction in modular polyketide synthases.
When an enoylreductase enzyme of a modular polyketide synthase reduces a propionate extender unit that has been newly added to the growing polyketide chain, the resulting methyl branch may have either S or R configuration. We have uncovered a correlation between the presence or absence of a unique tyrosine residue in the ER active site and the chirality of the methyl branch that is introduced. When this position in the active site is occupied by a tyrosine residue, the methyl branch has S configuration, otherwise it has R configuration. In a model PKS in vivo, a mutation (Tyr to Val) in an erythromycin PKS-derived ER caused a switch in the methyl branch configuration in the product from S to R. In contrast, alteration (Val to Tyr) at this position in a rapamycin-derived PKS ER was insufficient to achieve a switch from R to S, showing that additional residues also participate in stereocontrol of enoylreduction.
Kwan
David H
DH
Department of Biochemistry, University of Cambridge, 80 Tennis Court Road, Cambridge CB2 1GA, UK.
Sun
Yuhui
Y
Schulz
Frank
F
Hong
Hui
H
Popovic
Bojana
B
Sim-Stark
Joalice C C
JC
Haydock
Stephen F
SF
Leadlay
Peter F
PF
D000595:Amino Acid Sequence; D020134:Catalytic Domain; D002473:Cell Wall / Q000378:metabolism; D005227:Fatty Acids / Q000378:metabolism; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009161:Mycobacterium / Q000166:cytology; D044925:Oxidoreductases Acting on CH-CH Group Donors / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D048630:Polyketide Synthases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D011422:Propionates / Q000378:metabolism; D015202:Protein Engineering / Q000379:methods*; D017434:Protein Structure, Tertiary; D016930:Saccharopolyspora / Q000201:enzymology*; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013237:Stereoisomerism
D005227:Fatty Acids; D011422:Propionates; D048630:Polyketide Synthases; D044925:Oxidoreductases Acting on CH-CH Group Donors; C083835:2-methyl branched-chain enoyl-CoA reductase