Insights into bacterial 6-methylsalicylic acid synthase and its engineering to orsellinic acid synthase for spirotetronate generation.
The enzymes 6-methylsalicylic acid (6-MSA) synthases (6-MSASs) are involved in the building of an aryl moiety in many bioactive secondary metabolites produced by fungi and bacteria. Using the bacterial 6-MSAS ChlB1 in the biosynthesis of spirotetronate antibiotic chlorothricin (CHL) as a model, functional analysis of its dehydratase (DH) and ketoreductase (KR) domains by site-specific mutagenesis revealed that selective ketoreduction is not essential for polyketide chain extension. Promiscuity of the ketoacylsynthase domain in beta functionality recognition allows for engineering ChlB1 to an orsellinic acid (OSA) synthase (OSAS) by inactivating KR at the active site. The engineered ChlB1 is compatible with the enzymes for late-stage tailoring in CHL biosynthesis, featuring specific protein recognitions that facilitate variable aryl group incorporation. The resulting spirotetronates, which bear an OSA-derived aryl group, exhibited antibacterial activities comparable to those of the parent products.
State Key Laboratory of Bioorganic and Natural Products Chemistry, Shanghai Institute of Organic Chemistry, Chinese Academy of Sciences, 345 Lingling Road, Shanghai 200032, China.
Wei
W
Ding
Chun
C
Lei
Qingli
Q
He
Qinglin
Q
Zhang
Yurong
Y
Bi
Wen
W
Liu
D000217:Acyltransferases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000617:Aminoglycosides / Q000096:biosynthesis* / Q000737:chemistry; D000900:Anti-Bacterial Agents / Q000096:biosynthesis* / Q000737:chemistry; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D008025:Ligases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D009097:Multienzyme Complexes / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D010084:Oxidation-Reduction; D010088:Oxidoreductases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015202:Protein Engineering; D017434:Protein Structure, Tertiary; D012118:Resorcinols / Q000378:metabolism; D012459:Salicylates / Q000378:metabolism; D013302:Streptomyces / Q000201:enzymology
D000617:Aminoglycosides; D000900:Anti-Bacterial Agents; D001426:Bacterial Proteins; D009097:Multienzyme Complexes; D012118:Resorcinols; D012459:Salicylates; C501612:orsellinic acid; D010088:Oxidoreductases; D000217:Acyltransferases; C021587:6-methylsalicylic acid synthetase; D008025:Ligases; C000451:chlorothricin; C005236:6-methylsalicylic acid