1HL2

Crystal structure of N-acetylneuraminate lyase from E. coli mutant L142R in complex with b-hydroxypyruvate

1HL2 の概要

• エントリーDOI: 10.2210/pdb1hl2/pdb
• 関連するPDBエントリー: 1FDY 1FDZ 1NAL
• 分子名称: N-ACETYLNEURAMINATE LYASE SUBUNIT, 3-HYDROXYPYRUVIC ACID (3 entities in total)
• 機能のキーワード: n-acetylneuraminate lyase, class i aldolase, lyase, carbohydrate metabolism, schiff base
• 由来する生物種: ESCHERICHIA COLI
• タンパク質・核酸の鎖数: 4
• 化学式量合計: 131097.88

構造登録者

Joerger, A.C.,Fersht, A.R. (登録日: 2003-03-12, 公開日: 2003-05-09, 最終更新日: 2024-11-06)

主引用文献

Joerger, A.C.,Mayer, S.,Fersht, A.R.
Mimicking Natural Evolution in Vitro: An N-Acetylneuraminate Lyase Mutant with an Increased Dihydrodipicolinate Synthase Activity
Proc.Natl.Acad.Sci.USA, 100:5694-, 2003

PubMed Abstract: 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.

PubMed: 12711733
DOI: 10.1073/PNAS.0531477100

実験手法

X-RAY DIFFRACTION (1.8 Å)