1H and 13C NMR spectroscopic studies of the ferriheme resonances of three low-spin complexes of wild-type nitrophorin 2 and nitrophorin 2V24E as a function of pHReportar como inadecuado




1H and 13C NMR spectroscopic studies of the ferriheme resonances of three low-spin complexes of wild-type nitrophorin 2 and nitrophorin 2V24E as a function of pH - Descarga este documento en PDF. Documentación en PDF para descargar gratis. Disponible también para leer online.

JBIC Journal of Biological Inorganic Chemistry

, Volume 14, Issue 7, pp 1077–1095

First Online: 11 June 2009Received: 16 March 2009Accepted: 24 May 2009

Abstract

The ferriheme resonances of the low-spin S = 1-2 complexes of wild-type wt nitrophorin 2 NP2 and its heme pocket mutant NP2V24E with imidazole ImH, histamine Hm, and cyanide CN as the sixth ligand have been investigated by NMR spectroscopy as a function of pH 4.0–7.5. For the three wt NP2 complexes, the ratio of the two possible heme orientational isomers, A and B , remains almost unchanged ratio of A :B approximately 1:6 to 1:5 over this wide pH range. However, strong chemical exchange cross peaks appear in the nuclear Overhauser effect spectroscopy-exchange spectroscopy NOESY-EXSY spectra for the heme methyl resonances at low pH pH* 4.0–5.5, which indicate chemical exchange between two species. We have shown these to be two different exogenous ImH or Hm orientations that are denoted B and B ′, with the ImH plane nearly parallel and perpendicular to the ImH plane of the protein-provided His57, respectively. The wt NP2–CN complex also shows EXSY cross peaks due to chemical exchange, which is shown to be a result of interchange between two ruffling distortions of the heme. The same ruffling distortion interchange is also responsible for the ImH and Hm chemical exchange. For the three NP2V24E ligand complexes, no EXSY cross peaks are observed, but the A :B ratios change dramatically with pH. The fact that heme favors the A orientation highly for NP2V24E at low pH as compared with wt NP2 is believed to be due to the steric effect of the V24E mutation. The existence of the B ′ species at lower pH for wt NP2 complexes and the increase in A heme orientation at lower pH for NP2V24E are believed to be a result of a change in structure near Glu53 when it is protonated at low pH. H{C} heteronuclear multiple quantum coherence HMQC spectra are very helpful for the assignment of heme and nearby protein side chain resonances.

KeywordsNitrophorin NMR Ferriheme Glutamate mutant Imidazole complex Electronic supplementary materialThe online version of this article doi:10.1007-s00775-009-0551-3 contains supplementary material, which is available to authorized users.

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Autor: Fei Yang - Markus Knipp - Tatiana K. Shokhireva - Robert E. Berry - Hongjun Zhang - F. Ann Walker

Fuente: https://link.springer.com/







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