Amino-acid sequence of the cytochrome-b5-like heme-binding domain from Hansenula anomala flavocytochrome b2.
Flavocytochrome b2 (L-lactate dehydrogenase) from baker's yeast is composed of two structural and functional domains. Its first 100 residues constitute the heme-binding core, which is homologous to cytochrome b5 [B. Guiard, O. Groudinsky & F. Lederer (1974) Proc. Natl Acad. Sci. USA 71, 2539-2543]. We report here the amino acid sequence of the heme-binding domain isolated by tryptic proteolysis of Hansenula anomala flavocytochrome b2. The sequence was established by automated degradation of the whole fragment and of peptides obtained by CNBr cleavage at the unique tryptophan and by proteolysis with thermolysin and endoproteinase Lys C. As isolated, the domain consists of 84 residues without any sulfur amino acids. It shows 49 identities with the heme-binding domain from Saccharomyces cerevisiae and 28 with beef microsomal cytochrome b5. Using the recently published three-dimensional structure of S. cerevisiae flavocytochrome b2 [Z-x. Xia, N. Shamala, P. H. Bethge, L. W. Lim, H. D. Bellamy, N. H. Xuong, F. Lederer and F. S. Mathews (1987) Proc. Natl Acad. Sci. USA 84, 2629-2633], it can be seen that there are only positively charged side chains close to the accessible heme edge, the only negative charges in that area being those of the heme propionates. The implications of this result are discussed in the light of Salemme's model for the cytochrome b5/cytochrome c complex [F. R. Salemme (1976) J. Mol. Biol. 102, 563-568].
Haumont
P Y
PY
Centre National de la Recherche Scientifique Unité Associée 122, Hôpital Necker, Paris, France.
Thomas
M A
MA
Labeyrie
F
F
Lederer
F
F
D000595:Amino Acid Sequence; D000818:Animals; D001665:Binding Sites; D002417:Cattle; D003573:Cytochrome b Group / Q000032:analysis*; D015786:Cytochromes b5; D006418:Heme / Q000378:metabolism*; D007770:L-Lactate Dehydrogenase / Q000032:analysis*; D042844:L-Lactate Dehydrogenase (Cytochrome); D008862:Microsomes, Liver / Q000201:enzymology; D008969:Molecular Sequence Data; D010843:Pichia / Q000201:enzymology*; D012441:Saccharomyces cerevisiae / Q000201:enzymology; D004718:Saccharomycetales / Q000201:enzymology*
D003573:Cytochrome b Group; D006418:Heme; D015786:Cytochromes b5; D007770:L-Lactate Dehydrogenase; D042844:L-Lactate Dehydrogenase (Cytochrome)