Preparation of a whole-cell biocatalyst of mutated Candida antarctica lipase B (mCALB) by a yeast molecular display system and its practical properties.
To prepare a whole-cell biocatalyst of a stable lipase at a low price, mutated Candida antarctica lipase B (mCALB) constructed on the basis of the primary sequences of CALBs from C. antarctica CBS 6678 strain and from C. antarctica LF 058 strain was displayed on a yeast cell surface by alpha-agglutinin as the anchor protein for easy handling and stability of the enzyme. When mCALB was displayed on the yeast cell surface, it showed a preference for short chain fatty acids, an advantage for producing flavors; although when Rhizopus oryzae lipase (ROL) was displayed, the substrate specificity was for middle chain lengths. When the thermal stability of mCALB on the cell surface was compared with that of ROL on a cell surface, T (1/2), the temperature required to give a residual activity of 50% for heat treatment of 30 min, was 60 degrees C for mCALB and 44 degrees C for ROL indicating that mCALB displayed on cell surface has a higher thermal stability. Furthermore, the activity of the displayed mCALB against p-nitrophenyl butyrate was 25-fold higher than that of soluble CALB, as reported previously. These findings suggest that mCALB-displaying yeast is more practical for industrial use as the whole-cell biocatalyst.
Kato
Michiko
M
Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Sakyo-ku, Kyoto, 606-8502, Japan. mk@kais.kyoto-u.ac.jp
Fuchimoto
Jun
J
Tanino
Takanori
T
Kondo
Akihiko
A
Fukuda
Hideki
H
Ueda
Mitsuyoshi
M
D002175:Candida / Q000201:enzymology* / Q000235:genetics; D002473:Cell Wall / Q000201:enzymology; D004795:Enzyme Stability; D005656:Fungal Proteins; D008049:Lipase / Q000235:genetics / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D008858:Microscopy, Phase-Contrast; D009154:Mutation; D012233:Rhizopus / Q000201:enzymology; D013379:Substrate Specificity; D013696:Temperature
D005656:Fungal Proteins; D008049:Lipase; C431349:lipase B, Candida antarctica