Conformational changes and activity alterations induced by nickel ion in horseradish peroxidase.

Abstract:

:Conformational changes induced by the binding of nickel to horseradish peroxidase C (HRPC) were studied by electronic absorption spectroscopy, fluorescence spectroscopy and circular dichroism spectroscopy. Incubation of HRPC with various concentrations of Ni(2+) for 5 minutes resulted in changes in the enzyme absorption spectrum, including variations in the intensities of the Soret, beta and charge transfer (CT1) bands absorption, shift in the Soret, beta and CT1 bands maxima and absorption increase at 275 nm. Increases in the enzyme's intrinsic fluorescence as determined by fluorescence spectroscopy, as well as changes in the alpha-helical content, as determined by circular dichroism spectroscopy, were also found. Correlatively, alterations of the enzymatic activity by Ni(2+) were studied by following the H(2)O(2)-mediated oxidation of o-dianisidine and 2,2'-azinobis(3-ethylbenzothiazolinesulfonic acid) (ABTS) by HRPC. With both reducing substrates, it was found that in the presence of sufficient amount of enzyme, 1-10 mM nickel would enhance the enzymatic activity, while higher Ni(2+) concentrations (20-50 mM) would inhibit it. The enzyme was completely inhibited after 5 minutes incubation in 50 mM Ni(2+). Prolonged incubation would induce complete inhibition at lower Ni(2+) concentrations. Spectrophotometry investigations also showed that inhibitory concentrations of Ni(2+) altered compounds I and II formation, compound II being the first affected. Based on spectrophotometry, fluorescence and circular dichroism spectroscopy, and data on compounds I and II formation, a scheme is suggested for HRPC conformational changes in different Ni(2+) concentrations. HRPC was found to have four potential attachment sites for Ni(2+) which were sequentially occupied in a dose- and time-dependent manner by the metallic ion.

journal_name

Biochimie

journal_title

Biochimie

authors

Tayefi-Nasrabadi H,Keyhani E,Keyhani J

doi

10.1016/j.biochi.2006.04.001

subject

Has Abstract

pub_date

2006-09-01 00:00:00

pages

1183-97

issue

9

eissn

0300-9084

issn

1638-6183

pii

S0300-9084(06)00051-4

journal_volume

88

pub_type

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