The quinohaemoprotein lupanine hydroxylase from Pseudomonas putida

被引:9
作者
Hopper, DJ [1 ]
Kaderbhai, MA [1 ]
机构
[1] Univ Wales, Inst Biol Sci, Aberystwyth SY23 3DD, Ceredigion, Wales
来源
BIOCHIMICA ET BIOPHYSICA ACTA-PROTEINS AND PROTEOMICS | 2003年 / 1647卷 / 1-2期
关键词
quinohaemoprotein; cytochrome; PQQ; renaturation; reactivation; lupanine;
D O I
10.1016/S1570-9639(03)00070-0
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Lupanine hydroxylase catalyses the first reaction in the catabolism of the alkaloid lupanine by Pseudomonas putida. It dehydrogenates the substrate, which can then be hydrated. It is a monomeric protein of M, 72,000 and contains a covalently bound haem and a molecule of PQQ. The gene for this enzyme has been cloned and sequenced and the derived protein sequence has a 26 amino acid signal sequence at the N-terminal for translocation of the protein to the periplasm. Many of the features seen in the sequence of lupanine hydroxylase are common with other quinoproteins including the W-motifs that are characteristic of the eight-bladed propeller structure of methanol dehydrogenase. However, the unusual disulfide bridge between adjacent cysteines that is present in some PQQ-containing enzymes is absent in lupanine hydroxylase. The C-terminal domain contains characteristics of a cytochrome e and overall the sequence shows similarities with that of the quinohaemoprotein, alcohol dehydrogenase from Comamonas testosteroni. The gene coding for lupanine hydroxylase has been successfully expressed in Escherichia coli and a procedure has been developed to renature and reactivate the enzyme, which was found to be associated with the inclusion bodies. Reactivation required addition of PQQ and was dependent on calcium ions. (C) 2003 Elsevier Science B.V. All rights reserved.
引用
收藏
页码:110 / 115
页数:6
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