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Engineering copper sites in proteins: Loops confer native structures and properties to chimeric cupredoxins

Lookup NU author(s): Dr Chan Li, Dr Mark Banfield, Professor Christopher Dennison

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Abstract

The ligand-containing loops of two copper-binding electron-transfer proteins (cupredoxins) have been swapped. In the azurin (AZ) variant in which the plastocyanin (PC) sequence is introduced (AZPC), the loop adopts a conformation identical to that in PC. The reduction potential of AZPC is raised as compared to AZ and matches that of PC. In the previously published AZAMI variant (AMI = amicyanin), the shorter introduced loop adopts the same conformation as in AMI, and the reduction potential is lowered to equal that of AMI (Yanagisawa, S.; Dennison, C. J. Am. Chem. Soc. 2004, 126, 15711-15719. Li, C.; et al. Proc. Natl. Acad. Sci. U.S.A. 2006, 103, 7258-7263). Thus, the loop structure plays an important role in tuning the reduction potential of a type 1 copper site with contributions from protein dipoles in this region probably the most important feature. The structure of the loop also seems to be a major factor in controlling dissociation and protonation of the C-terminal His ligand, which can act as a switch to regulate electron-transfer reactivity. The PCAZ variant (PC with the AZ loop) possesses an active site, which is different from those of both PC and AZ, and it is assumed that the introduced loop does not adopt a structure as in AZ. This contributes to the observed instability of PCAZ and highlights that loop-scaffold interactions are important for stabilizing the active site of a cupredoxin. © 2007 American Chemical Society.


Publication metadata

Author(s): Li C, Banfield MJ, Dennison C

Publication type: Article

Publication status: Published

Journal: Journal of the American Chemical Society

Year: 2007

Volume: 129

Issue: 3

Pages: 709-718

ISSN (print): 0002-7863

ISSN (electronic): 1943-2984

Publisher: American Chemical Society

URL: http://dx.doi.org/10.1021/ja0661562

DOI: 10.1021/ja0661562

PubMed id: 17227035


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Funding

Funder referenceFunder name
BB/C504519/1Biotechnology and Biological Sciences Research Council

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