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'''CRYSTAL STRUCTURE OF N-ACETYLNEURAMINATE LYASE FROM E. COLI MUTANT L142R IN COMPLEX WITH B-HYDROXYPYRUVATE''' | '''CRYSTAL STRUCTURE OF N-ACETYLNEURAMINATE LYASE FROM E. COLI MUTANT L142R IN COMPLEX WITH B-HYDROXYPYRUVATE''' | ||
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[[Category: Fersht, A R.]] | [[Category: Fersht, A R.]] | ||
[[Category: Joerger, A C.]] | [[Category: Joerger, A C.]] | ||
[[Category: | [[Category: Carbohydrate metabolism,schiff base]] | ||
[[Category: | [[Category: Class i aldolase,lyase]] | ||
[[Category: | [[Category: N-acetylneuraminate lyase]] | ||
''Page seeded by [http://oca.weizmann.ac.il/oca OCA ] on Fri May 2 18:58:04 2008'' | |||
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Revision as of 18:58, 2 May 2008
CRYSTAL STRUCTURE OF N-ACETYLNEURAMINATE LYASE FROM E. COLI MUTANT L142R IN COMPLEX WITH B-HYDROXYPYRUVATE
OverviewOverview
N-acetylneuraminate lyase (NAL) and dihydrodipicolinate synthase (DHDPS) belong to the NAL subfamily of (betaalpha)(8)-barrels. They share a common catalytic step but catalyze reactions in different biological pathways. By rational design, we have introduced various mutations into the NAL scaffold from Escherichia coli to switch the activity toward DHDPS. These mutants were tested with respect to their catalytic properties in vivo and in vitro as well as their stability. One point mutation (L142R) was sufficient to create an enzyme that could complement a bacterial auxotroph lacking the gene for DHDPS as efficiently as DHDPS itself. In vitro, this mutant had an increased DHDPS activity of up to 19-fold as defined by the specificity constant k(cat)K(M) for the new substrate l-aspartate-beta-semialdehyde when compared with the residual activity of NAL wild-type, mainly because of an increased turnover rate. At the same time, mutant L142R maintained much of its original NAL activity. We have solved the crystal structure of mutant L142R at 1.8 A resolution in complex with the inhibitor beta-hydroxypyruvate. This structure reveals that the conformations of neighboring active site residues are left virtually unchanged by the mutation. The high flexibility of R142 may favor its role in assisting in catalysis. Perhaps, nature has exploited the catalytic promiscuity of many enzymes to evolve novel enzymes or biological pathways during the course of evolution.
About this StructureAbout this Structure
1HL2 is a Single protein structure of sequence from Escherichia coli. Full crystallographic information is available from OCA.
ReferenceReference
Mimicking natural evolution in vitro: an N-acetylneuraminate lyase mutant with an increased dihydrodipicolinate synthase activity., Joerger AC, Mayer S, Fersht AR, Proc Natl Acad Sci U S A. 2003 May 13;100(10):5694-9. Epub 2003 Apr 23. PMID:12711733 Page seeded by OCA on Fri May 2 18:58:04 2008