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

Abstract

Heme degradation reaction of HutZ from Vibrio cholerae is regulated by the subunit–subunit interactions.

Role of conserved arginine in the heme distal site of HutZ from Vibrio cholerae in the heme degradation reaction

“…Recent studies by our group further showed that subunit-subunit interactions play a key role in the heme degradation reaction [16]. The distance between heme and Trp109, which reflects subunit-subunit orientational changes, was increased from [16][17][18] Å for wild-type HutZ to 23-25 Å for R92A and R92L mutant enzymes, similar to data obtained for mutants of Ala31 [16], His63 [19] and Asp132. Based on the collective results, we propose an activation mechanism of HutZ involving conformational changes at Arg92, which are transmitted to the heme axial ligand His170 through Ala31, His63, and Asp132, with modulation of subunit-subunit interactions.…”

“…Heme degradation activity was also pH-dependent; HutZ is active at pH 6.0, but it is inactive at pH >7.0 [16]. In view of the correlation between the high-spin heme content and heme degradation activity, we propose that heme degradation activity of HutZ is related to the high-spin heme content [16].…”

“…Accordingly, almost no heme degradation occurs in heme-HutZ at pH values of 7.0 or above [16]. In contrast, the heme iron at pH 6.0 is reduced faster relative to that under neutral or alkaline conditions by ~3-fold, potentially due to lower electron donation from proximal histidine [16]. Thus, heme in the heme-HutZ complex is efficiently degraded at pH 6.0 but not pH >7.0.…”

“…The half-life of the oxyferrous heme of HutZ at pH 8.0 is less than 10 s [15], whereas that of heme-HO is more than 1 h [17]. Accordingly, almost no heme degradation occurs in heme-HutZ at pH values of 7.0 or above [16]. In contrast, the heme iron at pH 6.0 is reduced faster relative to that under neutral or alkaline conditions by ~3-fold, potentially due to lower electron donation from proximal histidine [16].…”

“…The plots of Fe-CO (ν Fe-CO ) and C-O (ν C-O ) stretching modes of the CO-bound form are far off from the line with proteins possessing neutral histidine as an axial ligand but near the line with proteins with imidazolate as the axial ligand. The imidazolate character of the proximal histidine with negative redox potential of heme from heme-HutZ at pH 8.0 (-176 mV) leads to stabilization of ferric rather than ferrous heme, resulting in a slow reduction rate of heme [16]. In addition, even if heme is reduced, oxyferrous heme is rapidly re-oxidized to ferric heme via autoxidation.…”