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Title: Para-nitrobenzyl esterases with enhanced activity in aqueous and nonaqueous media

Abstract

A method is disclosed for isolating and identifying modified para-nitrobenzyl esterases which exhibit improved stability and/or esterase hydrolysis activity toward selected substrates and under selected reaction conditions relative to the unmodified para-nitrobenzyl esterase. The method involves preparing a library of modified para-nitrobenzyl esterase nucleic acid segments (genes) which have nucleotide sequences that differ from the nucleic acid segment which encodes for unmodified para-nitrobenzyl esterase. The library of modified para-nitrobenzyl nucleic acid segments is expressed to provide a plurality of modified enzymes. The clones expressing modified enzymes are then screened to identify which enzymes have improved esterase activity by measuring the ability of the enzymes to hydrolyze the selected substrate under the selected reaction conditions. Specific modified para-nitrobenzyl esterases are disclosed which have improved stability and/or ester hydrolysis activity in aqueous or aqueous-organic media relative to the stability and/or ester hydrolysis activity of unmodified naturally occurring para-nitrobenzyl esterase. 43 figs.

Inventors:
;
Publication Date:
Research Org.:
California Institute of Technology
Sponsoring Org.:
USN; USDOE, Washington, DC (United States); Department of the Navy, Washington, DC (United States)
OSTI Identifier:
6479844
Patent Number(s):
US 5906930; A
Application Number:
CNN: N00014-91-J-1397; PPN: US 9-020991
Assignee:
California Inst. of Tech., Pasadena, CA (United States) CHO; EDB-99-081942
DOE Contract Number:  
FG02-93CH10578
Resource Type:
Patent
Resource Relation:
Patent File Date: 9 Feb 1998
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES; ENZYMATIC HYDROLYSIS; ENZYME ACTIVITY; ESTERASES; ESTERS; GENES; CHEMICAL REACTIONS; DECOMPOSITION; ENZYMES; HYDROLASES; HYDROLYSIS; LYSIS; ORGANIC COMPOUNDS; PROTEINS; SOLVOLYSIS; 550200* - Biochemistry

Citation Formats

Arnold, F.H., and Moore, J.C. Para-nitrobenzyl esterases with enhanced activity in aqueous and nonaqueous media. United States: N. p., 1999. Web.
Arnold, F.H., & Moore, J.C. Para-nitrobenzyl esterases with enhanced activity in aqueous and nonaqueous media. United States.
Arnold, F.H., and Moore, J.C. Tue . "Para-nitrobenzyl esterases with enhanced activity in aqueous and nonaqueous media". United States.
@article{osti_6479844,
title = {Para-nitrobenzyl esterases with enhanced activity in aqueous and nonaqueous media},
author = {Arnold, F.H. and Moore, J.C.},
abstractNote = {A method is disclosed for isolating and identifying modified para-nitrobenzyl esterases which exhibit improved stability and/or esterase hydrolysis activity toward selected substrates and under selected reaction conditions relative to the unmodified para-nitrobenzyl esterase. The method involves preparing a library of modified para-nitrobenzyl esterase nucleic acid segments (genes) which have nucleotide sequences that differ from the nucleic acid segment which encodes for unmodified para-nitrobenzyl esterase. The library of modified para-nitrobenzyl nucleic acid segments is expressed to provide a plurality of modified enzymes. The clones expressing modified enzymes are then screened to identify which enzymes have improved esterase activity by measuring the ability of the enzymes to hydrolyze the selected substrate under the selected reaction conditions. Specific modified para-nitrobenzyl esterases are disclosed which have improved stability and/or ester hydrolysis activity in aqueous or aqueous-organic media relative to the stability and/or ester hydrolysis activity of unmodified naturally occurring para-nitrobenzyl esterase. 43 figs.},
doi = {},
journal = {},
number = ,
volume = ,
place = {United States},
year = {Tue May 25 00:00:00 EDT 1999},
month = {Tue May 25 00:00:00 EDT 1999}
}