The N-Terminal Amino Acid Residues of Normal Adult Human Hemoglobin: A Quantitative Study of Certain Aspects of Sanger's DNP-Method

Abstract

A quantitative redetermination of the N-terminal valyl residues of normal adult human hemoglobin by the DNP-method of Sanger has led us to question the validity of results previously reported. Our experimental results indicate that there are 3.6 N-terminal valyl residues per molecule, based on a molecular weight of 66,700 for human hemoglobin. The essential difference between this value and those of other investigators lies in a correction factor for operational, chromatographic and hydrolytic losses (13%) which is appreciably lower than any previously reported value. This low value is justified by a detailed study of losses with DNP-valine and two peptides, DNP-Val-gly and DNP-Val-leu, the latter an important hydrolytic product of human hemoglobin itself. On the basis of these results an integral value for the number of end groups in human hemoglobin can be achieved only by revising the molecular weight, If, on the other hand, the number of N-terminal valyl residues in human hemoglobin is non-integral, it may well indicate that normal adult human hemoglobin contains more than one kind of molecule.

A New N-Terminal Blocking Group Involving a Schiff Base in Hemoglobin AIc*

“…and in j3f3Alc'Globin by Sanger's Procedure. The experimental procedures for dinitrophenylation and the isolation of N-terminal DNP peptides and DNP-amino acids followed Rhinesmith et al (1957aRhinesmith et al ( ,b, 1958.…”

Chemical Characterization and Subunit Hybridization of Human Hemoglobin H and Associated Compounds*

“…the a chain, DNP-val-his-leu and DNP-val-his from the 0 chain, and DNP-glycine from the 7 chain; and (3) that, in long hydrolyses, DNP-valine arises from the 0 chain. The basis for these assumptions has been described elsewhere (Rhinesmith et al, 1957a(Rhinesmith et al, , 1958Schroeder and Matsuda, 1958). The number of chains of each type in a given molecule is then obtained by summation of the appropriate figures as presented in the right-hand three columns of Table V.…”