¹H and ¹³C NMR Studies of a Truncated Heme Domain from Chlorella vulgaris Nitrate Reductase:
Signal Assignment of the Heme Moiety
 
Xiangdong Wei and Li-June Ming*
Department of Chemistry and Institute for Biomolecular Science
University of South Florida, Tampa, Florida 33620-5250

Andrew C. Cannons
Department of Biology and Institute for Biomolecular Science
University of South Florida, Tampa, Florida 33612

Larry P. Solomonson
Department of Biochemistry and Molecular Biology, College of Medicine
and Institute for Biomolecular Science
University of South Florida, Tampa, Florida 33612

Running Title:  NMR of Chlorella Nitrate Reductase Heme-Domain
Key Words:  2D NMR; heme proteins; cytochrome b; Nitrate reductase; Chlorella

Abstract:
A water soluble truncated heme domain (a tetramer of MW = 45 kDa) of the tetrameric nitrate reductase complex from the green alga Chlorella vulgaris has been overexpressed and purified.  This truncated heme domain with four identical subunits has a high redox potential (midpoint potential E1/2 = +16 mV) as compared with other heme-containing flavoproteins.  We have undertaken a determination of the detailed configuration of the heme moiety in order to understand the unique electrochemical property of the heme moiety of this enzyme.  We report here the study of the heme prosthetic group of the truncated heme domain by the use of 2D ¹H and ¹³C NMR techniques.  A complete signal assignment of the heme has been achieved. Our observations suggest that the heme configuration is similar to that of the crystal structure of the membrane-bound bovine liver cytochrome b5.
 
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