5 Molecular Basis of Specificity and
Stereoselectivity of Microbial Lipases
toward Triacylglycerols
Ju ̈rgen Pleiss
5.1 Biochemical properties
Microbial lipases are widely used to catalyze hydrolysis, alcoholysis, esterification
and transesterification of triacylglyerols and analogs, thereby taking advantage of
their catalytic efficiency, fatty acid specificity, regio- and stereoselectivity (for re-
views, see Faber, 1997; Schmid and Verger, 1998; Bornscheuer and Kazlauskas,
1999). Lipases are long-chain triglyceride-hydrolyzing enzymes, thereby being dis-
tinguished from esterases which hydrolyze soluble esters. They have a higher activ-
ity toward insoluble substrates than toward the same substrates in monomeric form,
an effect which has been termed ‘interfacial activation’ (Sarda and Desnuelle, 1958).
Lipase activity, specificity and selectivity depend upon reaction conditions such as
the quality of the substrate interface (Verger, 1997), immobilization support (Ad-
lercreutz, 1991), solvent (water or organic solvent for hydrolysis or esterification
reactions, respectively), and water activity in organic solvent (Halling, 1994). How-
ever, experimental set-ups which measure competitive factorsain mixtures of dif-
ferent fatty acid chains in organic solvent (Deleuze et al., 1987; Rangheard et al.,
1989; Borgdorf and Warwel, 1999) suppress these secondary effects. Thus, the com-
petitive factorawhich corresponds to the ratio of the specificity constantskcat/Kmis
independent of reaction conditions.
All lipases convert esters of medium (C 4 ) to long chain (C 18 ) saturated fatty acids.
Some of them efficiently hydrolyze fatty acid esters as long as C 22 , such as lipase
fromRhizomucor miehei(Rangheard et al., 1992; Kirk et al., 1992). Of special inter-
est is thecis(D-9) specificity ofGeotrichum candidumlipase I (Phillips et al., 1995;
Borgdorf and Warwel, 1999).
All lipases hydrolyze triacylglycerols at thesn-1/3 position; hydrolysis at thesn- 2
position is catalyzed only by four microbial lipases (Candida antarcticalipase A,
Geotrichum candidumlipase I,Penicillium simplicissimumlipase, andCandida
rugosalipase). Regioselectivity andsn-1/3 stereoselectivity were measured using
monomolecular films of di- and triacylgycerol analogues (Ransac et al., 1990), pro-
chiral triacylglycerol emulsions (Rogalska et al., 1993; Stadler et al., 1995; Kovac et
al., 1996) and optically pure diacylglycerols (Rogalska et al., 1995). Three lipases
were highly stereoselective toward trioctanoin: namely, lipases fromPseudomonas
sp. andPseudomonas aeruginosa(sn-1 selective), andCandida antarcticalipase B
(sn-3 selective). All other lipases show medium or low selectivity. Stereopreference
was similar toward both tri- and diacylglycerol substrates, and generally did not
depend on the type of substrate. Only forCandida antarcticalipase B andGeotri-
Enzymesin LipidModification.Editedby UweT. Bornscheuer
Copyright 2000 Wiley-VCHVerlagGmbH& Co. KGaA,Weinheim.ISBN:3-527-30176-3
