A heme- and metal-binding hexapeptide from the sequence of rabbit plasma histidine-rich glycoprotein.
Rabbit histidine-rich glycoprotein (HRG) binds low-spin heme and metals tightly at several sites that contain histidine. As part of an on-going effort to define and locate the binding sites for these and the other ligands of HRG, the sequence: NH2-Gly-His-Phe-Pro-Phe-His-Trp-... was found in a 16 kDa heme-binding peptide isolated from HRG. The spacing of the histidyl residues in this peptide, which contains the C-terminal 79 residues of HRG, together with molecular modeling suggested that this sequence might constitute one heme binding site of HRG by accommodating heme in a bis-histidyl linkage. Three peptides based on this sequence (I, HFPFHW; II, WHFPFH; and III, HFGFHW) were synthesized, and their ability to bind heme and metals examined. All three peptides bind heme as demonstrated by the changes produced in the absorbance of heme when mixed with the peptides. Substituting glycine for proline in the central position or moving the location of the tryptophan did not affect heme binding. The apparent Kd's of the mesoheme/peptide I, II and III complexes are 75 +/- 25 microM, indicative of heme binding approximately 100 times less avid than the mesoheme/HRG complex (Kd ca. 1 microM), but nearly 1000 times tighter than that of the mesoheme/histidine complex (Kd ca. 60 mM). The absorbance spectra of the mesoheme/peptide complexes, the loss of binding caused by modification of histidine residues, and the pH dependence of heme binding, all indicate that heme forms a low spin, bis-histidyl type of complex with these peptides, like that formed with HRG itself. Copper, but not cadmium or nickel, was an effective inhibitor of heme binding by the peptides. The sequence of HRG congruent with the sequence of peptide I is proposed to be one heme- and metal-binding site of rabbit HRG.
Morgan
W T
WT
Department of Biochemistry and Molecular Biology, LSU Medical Center, New Orleans 70112.
Deaciuc
V
V
Riehm
J P
JP
D000595:Amino Acid Sequence; D000818:Animals; D055598:Chemical Phenomena; D002621:Chemistry; D002851:Chromatography, High Pressure Liquid; D006023:Glycoproteins / Q000097:blood / Q000378:metabolism; D006418:Heme; D006863:Hydrogen-Ion Concentration; D008670:Metals / Q000378:metabolism*; D008969:Molecular Sequence Data; D011506:Proteins; D011817:Rabbits; D000099053:Histidine-Rich Glycoprotein
D006023:Glycoproteins; D008670:Metals; D011506:Proteins; D000099053:Histidine-Rich Glycoprotein; D006418:Heme