Structural significance of the amino-terminal residues of sperm whale myoglobin.
Following the development of a nondestructive synthetic procedure for rapid production of des-Val1-myoglobin in large quantities, the synthesis of a series of myoglobin derivatives varying in structure and charge in the NH2-terminal region was accomplished. In comparison to the untreated myoglobin, the des-Val1-myoglobin was found to possess at low pH a decreased stability and an increased net positive charge in the pH range 5.5-8.5. While the elevated net positive charge was no longer apparent after removal of the second residue, the instability of the molecule was found to be sharply increased. Substitutions of the first residue, directed toward elucidating its structural importance, included glutamic acid, lysine, and glycine. Addition of any of the three amino acids to the des-Val1-myoglobin was found to restore much of the acid stability, with the [Gly1]myoglobin appearing nearly identical with the native molecule. All three semisynthetic myoglobins showed potentiometric titration curves characteristic of their respective, substituted residue. Carbamylation of the NH2 terminal of myoglobin and des-Val1-myoglobin yielded two nearly identical molecules in terms of all physical properties examined. Consequently, it was concluded that the first residue primarily serves the function of maintaining the positively charged NH2 terminus a certain distance away from the beginning of the A helix and from the charge pair interaction of Lys-133 with Glu-6. In addition, through physical measurements of the des-Val1,Leu2-myoglobin prior and subsequent to carbamylation of the NH2 terminus, it was apparent that the stabilization conferred on the des-Val1-myoglobin by the second residue was dependent to a large degree upon the hydrophobic interactions of its side chain.
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002942:Circular Dichroism; D005998:Glycine; D006863:Hydrogen-Ion Concentration; D007202:Indicators and Reagents; D009211:Myoglobin; D011487:Protein Conformation; D013056:Spectrophotometry, Ultraviolet; D014633:Valine; D014907:Whales
D000596:Amino Acids; D007202:Indicators and Reagents; D009211:Myoglobin; D014633:Valine; D005998:Glycine