Gly311 residue triggers the enantioselectivity of Staphylococcus xylosus lipase: a monolayer study.
Using emulsified triacylglycerols, we have shown recently [Mosbah et al., 2007, submitted for publication] that amino acid residue G311 of Staphylococcus xylosus lipase (SXL) is critically involved in substrate selectivity, pH and temperature dependency. Using the monomolecular film technique, we show in the present study that the four single mutants of this residue (G311L, G311W, G311D, and G311K), interact efficiently with egg-phosphatidyl choline (egg-PC) monomolecular films, comparably to the wild-type (G311). A critical surface pressure (pi(c)) of about 25 mN/m was obtained with the SXL wild-type (SXL-WT) and its mutants. These results support our conclusion that the G311 residue is not involved in the interfacial adsorption step of SXL. A kinetic study on the surface pressure dependency, stereoselectivity, and regioselectivity of SXL-WT and its G311 mutants was also performed using optically pure enantiomers of diacylglycerols (1,2-sn-dicaprin and 2,3-sn-dicaprin) and a prochiral isomer (1,3-sn-dicaprin) spread as monomolecular films at the air-water interface. Our results indicated that the mutation of one single residue at position 311 affects critically the catalytic activity, the stereo- and the regioselectivity of SXL. As previously observed with emulsified substrates [Mosbah et al., 2007, submitted for publication] we observed that an increase in the size of the 311 amino acid side chain residue was accompanied by a decrease of lipase activity measured on dicaprin monolayer. We also noticed that the substitution of G311 by a basic or acidic residue (G311K and G311D), induces a significant shift of the pH optimum from 8 to 9.5 or from 8 to 6.5, respectively.
Mosbah
Habib
H
Laboratoire de biochimie et de génie enzymatique des lipases, ENIS BPW 3038 Sfax, Tunisia.
Sayari
Adel
A
Verger
Robert
R
Gargouri
Youssef
Y
D000595:Amino Acid Sequence; D005998:Glycine / Q000737:chemistry*; D006863:Hydrogen-Ion Concentration; D008049:Lipase / Q000737:chemistry* / Q000235:genetics; D010713:Phosphatidylcholines / Q000737:chemistry; D017354:Point Mutation; D013210:Staphylococcus / Q000201:enzymology; D013237:Stereoisomerism; D013499:Surface Properties; D053835:Unilamellar Liposomes / Q000737:chemistry*
D010713:Phosphatidylcholines; D053835:Unilamellar Liposomes; D008049:Lipase; D005998:Glycine