Autolytic site mutant R105C of rat trypsin.
The autolytic site Arg105 of rat trypsin was replaced with Cys by DNA site-directed mutagenesis method. Comparison of expression and purification of R105C trypsin along with the wild type and some other Arg105 mutants indicates that R105C trypsin could be expressed as well but with a lower expression level. It is unexpected that R105C trypsin has no detectable activity toward trypsin substrate TAME, quite different from the wild type and other Arg105 mutants. Native gel electrophoresis analysis indicates that R105C trypsin has a similar mobility rate to that of wild type trypsin. FPLC also gives similar retaining time. The loss of activity of R105C trypsin may result from the conformational change around active site, but not the dimer formation.
Li
X F
XF
National Laboratory of Protein Engineering and Plant Genetic Engineering College of Life Sciences, Peking University, Beijing, P.R. China.
Tang
J G
JG
Zhang
L X
LX
D000818:Animals; D004789:Enzyme Activation / Q000235:genetics; D016297:Mutagenesis, Site-Directed; D009154:Mutation; D011487:Protein Conformation; D051381:Rats; D014357:Trypsin / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
D014357:Trypsin