428 CORRADO BAGLIONI 



i'Us:>ih1 luMi' only with reference to i)apcr or starch-gel electrophoresis at 

 pH 8.6. which are most commonly used. It should be pointed out that 

 the abnormal hemogloi)ins may show a charge difference with Hb-A 

 variable with the pH and the nature of the electroi)horetic medium. The 

 behavior of some abnormal hemoglobins in agar-gel electrophoresis is 

 completely different from their behavior in starch-gel electrophoresis at 

 similar pH's I see review by dratzer and Bcaven, 1961) ; i.e., Hb-Dp„njah 

 and II1)-E are not resolved from llh-A by agar-gel electrophoresis at pH 

 6.2. wiiile they are resolved by other types of electrophoresis and by 

 chromatography on ion exchangers. It seems rather difficult to explain 

 the electrophoretic and chromatographic behavior of the abnormal 

 hemoglobins in terms of charge difTerence with Hb-A only; interactions 

 at the level of the tertiary structure of the hemoglobin molecule may 

 possibly account for some of the discrepancies observed. The charge of 

 the amino acids involved in substitutions and their position in the 

 tridimensional configuration of the ])rotein determine the electrophoretic 

 and chromatographic behavior of the abnormal hemoglobins. 



1. The Distributio?} of Amino Acid Substitutions in Hb-A Variants 



By determining the composition and the sequence of the peptides 

 altered in alinormal hemoglobins, it has been j^ossible to locate the 

 residues substituted in the Hb-A variants along the sequences of the 

 Hb-A peptide chains. In this way one can draw a map of the chemical 

 changes caused by known mutations of the hemoglobin genes (Fig. 5). 

 This is not a genetic map, but since the amino acid sequence of the 

 peptide chains is probably parallel or colinear with the nucleotide 

 sequence of the corresponding genes, it may correspond to a genetic map 

 of the mutations in the hemoglobin gene. Intragenic recombination is a 

 rare event and there is no doubt that the chances of observing such an 

 event in humans are ridiculously small; thus, there is no hope to prove 

 the colinearity between the amino acid sequence of a hemoglobin chain 

 and the genetic fine structure of the corresponding gene by means of 

 recombination of alleles. Demonstration of colinearity between genetic 

 map and amino acid sequence is now being sought in more favorable 

 organisms, as in Escherichia coli, for the gene controlling the enzyme 

 trytophan synthetase (Helinski and Yanofsky, 1962) and phosphatase 

 (Garen et al., 1961) or in bacteriophage for the gene controlling lysozyme 

 (Dreyer. 1960). 



In Fig. 5 is shown schematically the position along the peptide 

 chains of Hb-A of the amino acids substituted in the abnormal hemo- 

 globins. There is no particular clustering of substitutions in any region 

 of the peptide chains. It is remarkable, however, that some residues 



