Fast carbon-carbon bond formation by a promiscuous lipase.
Lipase B from Candida antarctica was redesigned to catalyze the promiscuous reaction of carbon-carbon bond formation. Mutation of the catalytic serine to alanine afforded a mutant that catalyzed Michael additions of 1,3-dicarbonyls to alpha,beta-unsaturated carbonyl compounds at high specific rates, such as 4000 s-1. The enzyme-catalyzed Michael addition reaction followed saturation kinetics and showed substrate inhibition. The designed enzyme showed high rate enhancements with a catalytic proficiency higher than 108, which is on the same level as that observed for enzymes with native substrates.
Svedendahl
Maria
M
Department of Biochemistry, School of Biotechnology, Royal Institute of Technology (KTH), AlbaNova University Center, SE-106 91 Stockholm, Sweden.
Hult
Karl
K
Berglund
Per
P
D000171:Acrolein / Q000737:chemistry / Q000378:metabolism; D002175:Candida / Q000201:enzymology; D002384:Catalysis; D005656:Fungal Proteins / Q000737:chemistry / Q000378:metabolism; D007700:Kinetics; D008049:Lipase / Q000737:chemistry* / Q000378:metabolism*; D016297:Mutagenesis, Site-Directed; D010422:Pentanones / Q000737:chemistry / Q000378:metabolism; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D005656:Fungal Proteins; D010422:Pentanones; C008790:acetylacetone; D000171:Acrolein; D008049:Lipase; C431349:lipase B, Candida antarctica