title: Efficient and selective isotopic labeling of hemes to facilitate the study of multiheme proteins.
abstract: Specific isotopic labeling of hemes provides a unique opportunity to characterize the structure and function of heme-proteins. Unfortunately, current methods do not allow efficient labeling in high yields of multiheme cytochromes c, which are of great biotechnological interest. Here, a method for production of recombinant multiheme cytochromes c in Escherichia coli with isotopically labeled hemes is reported. A small tetraheme cytochrome of 12 kDa from Shewanella oneidensis MR-1 was used to demonstrate the method, achieving a production of 4 mg pure protein per liter. This method achieves, in a single step, efficient expression and incorporation of hemes isotopically labeled in specific atom positions adequate for spectroscopic characterization of these complex heme proteins. It is, furthermore, of general application to heme proteins, opening new possibilities for the characterization of this important class of proteins.
authors: Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Oeiras, Portugal.
authors: Bruno M
authors: BM
authors: Fonseca
authors: Department of Biochemistry and Molecular Biology, Pennsylvania State University, University Park, PA, USA.
authors: Ming
authors: M
authors: Tien
authors: Department of Chemistry, University of Kansas, Lawrence, KS, USA.
authors: Mario
authors: M
authors: Rivera
authors: Microbiology Group, Pacific Northwest National Laboratory, Richland, WA, USA.
authors: Liang
authors: L
authors: Shi
authors: Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa, Oeiras, Portugal.
authors: Ricardo O
authors: RO
authors: Louro
mesh_terms: D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D006420:Hemeproteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D007553:Isotope Labeling / Q000379:methods*; D046912:Multiprotein Complexes / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D019906:Nuclear Magnetic Resonance, Biomolecular; D011994:Recombinant Proteins / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D020592:Shewanella / Q000737:chemistry / Q000235:genetics
chemical_list: D001426:Bacterial Proteins; D006420:Hemeproteins; D046912:Multiprotein Complexes; D011994:Recombinant Proteins
keywords: hemes; multiheme cytochromes c; specific isotopic labeling; δ-aminolevulinic acid