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Issue 12, 2019
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Subunit–subunit interactions play a key role in the heme-degradation reaction of HutZ from Vibrio cholerae

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Abstract

HutZ, a dimeric protein, from Vibrio cholerae is a protein that catalyzes the oxygen-dependent degradation of heme. Interestingly, the ascorbic acid-supported heme-degradation activity of HutZ depends on pH: less than 10% of heme is degraded by HutZ at pH 8.0, but nearly 90% of heme is degraded at pH 6.0. We examined here pH-dependent conformational changes in HutZ using fluorescence spectroscopy. Trp109 is estimated to be located approximately 21 Å from heme and is present in a different subunit containing a heme axial ligand. Thus, we postulated that the distance between heme and Trp109 reflects subunit–subunit orientational changes. On the basis of resonance energy transfer from Trp109 to heme, we estimated the distance between heme and Trp109 to be approximately 17 Å at pH 8.0, while the distance increased by less than 2 Å at pH 6.0. We presumed that such changes led to a decrease in electron donation from the proximal histidine, resulting in enhancement of the heme-degradation activity. To confirm this scenario, we mutated Ala31, located at the dimer interface, to valine to alter the distance through the subunit–subunit interaction. The distance between heme and Trp109 for the A31V mutant was elongated to 24–27 Å. Although resonance Raman spectra and reduction rate of heme suggested that this mutation resulted in diminished electron donation from the heme axial ligand, ascorbic acid-supported heme-degradation activity was not observed. Based on our findings, it can be proposed that the relative positioning of two protomers is important in determining the heme degradation rate by HutZ.

Graphical abstract: Subunit–subunit interactions play a key role in the heme-degradation reaction of HutZ from Vibrio cholerae

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Publication details

The article was received on 09 Feb 2019, accepted on 25 Feb 2019 and first published on 25 Feb 2019


Article type: Paper
DOI: 10.1039/C9DT00604D
Dalton Trans., 2019,48, 3973-3983

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    Subunit–subunit interactions play a key role in the heme-degradation reaction of HutZ from Vibrio cholerae

    T. Uchida, K. Ota, Y. Sekine, N. Dojun and K. Ishimori, Dalton Trans., 2019, 48, 3973
    DOI: 10.1039/C9DT00604D

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