Walter C. Mahoney

Reversed-phase supports for the resolution of large denatured protein fragments☆

Large pore (much greater than 300 A), spherical partical silica stationary phases possessing either C18 or C8 hydrocarbon ligands out-performed small pore (60-100A), irregular shaped silicas for the purification of large denatured peptides. Since columns 5 cm in length appeared to be as effective in separating peptides as columns 5 times longer, it is likely that large peptides absorb to the matrix rather than partition between the stationary and mobile phases.

Comparison of the antigenic properties of the glycinin polypeptides

Abstract The antigenic properties of purified glycinin subunits were studied using antibodies prepared against them. Antisera against native glycinin did not react with the isolated subunits, and antibodies prepared against the purified subunits were not active against native glycinin. When native glycinin -was denatured, the antiglycinin immunoglobulins lost their ability to react with it, although the denatured complex was then recognized by antibodies against the purified subunits. Substantial structural rearrangement apparently occurred when the native complex was denatured and disaggregated. Acidic polypeptides A 1a , A 1b , and A 2 had similar determinants as judged by their reactions against A 1a and A 1a antisera. The reaction of the A 3 polypeptides with these antibodies was of lower intensity and in each case clear spurs of cross-reactivity were visible. No cross-reaction was detected between polypeptide A 4 and either anti-A 1a or A 2 . Anti-A 3 antibodies reacted with each of the acidic polypeptides of glycinin, and distinct spurs of cross-reactivity were observed between A 3 vs A 1a , A 3 vs A 2 , and A 3 vs A 4 . B 1 Antisera developed a reaction of identity between basic polypeptides B 1 and B 2 , but reacted very weakly with B 3 and B 4 . The acidic and basic polypeptides of glycinin were immunologically unrelated. The results demonstrated that immunological tests would successfully differentiate some members of the family of acidic subunits, and other immunoglobulins would discriminate between members of the family of basic subunits.

Complete Primary Structure of the 3 Chain From the Hemoglobin of a Baboon, Papio cynocephalus

The complete primary structure of the beta chain from the adult hemoglobin of a baboon, Papio cynocephalus, has been determined by automated, Edman degradation of the intact chain and four fragments derived therefrom by specific cleavage reactions. The analysis was facilitated by application of a modified solvent system that permits unambiguous identification, by high-performance liquid chromatography, of the 17 amino acids whose phenylthiohydantoin derivatives are soluble in ethyl acetate. The sequence obtained differs from that of the human beta chain at eight sites, a degree of divergence similar to that observed when human and macaque beta chains are compared. Of the cercopithecoid beta chains whose complete sequences have been determined or inferred from compositions of small peptides, that of P. cynocephalus is most like the beta chain of the gelada baboon, an observation in accord with assessment of a close phylogenetic relationship between the genera Papio and Theropithecus.

Hemoglobin α-chain variation in macaques: Primary structures of the αI and αII chains from the adult hemoglobins of Malaysian Macaca nemestrina

Abstract The complete primary structures of each of the two α chains commonly found in the adult hemoglobins of Malaysian Macaca nemestrina (pig-tailed macaques) were obtained from the intact chains and five fragments produced by two nonenzymatic and three enzymatic cleavage reactions. The two chains differ at a single site, the α I chains having a glutaminyl residue and the α II chains having a histidinyl residue in position 78. Both chains differ from their human counterpart at five positions, the extent of divergence being similar to that observed for most of the α chains from other species of Macaca that have been analyzed to date. Elucidation of the structural difference between the α I and α II chains demonstrates that the high degree of heterogeneity observed among the hemoglobin phenotypes of M. nemestrina is a consequence of underlying genetic variability and not a result of postsynthetic modification of genetically identical proteins. Comparisons with the hemoglobin phenotypes found in other species of Macaca support the contention that chromosomes bearing linked α I and α II genes, as well as those bearing single α I and α II alleles, combine in zygotes to produce the phenotypic variation observed in members of Malaysian populations of M. nemestrina .

Complete Sequence of the γ Chain from the Fetal Hemoglobin of the Baboon, Papio cynocephalus

The amino acid sequence of the hemoglobin γ chain from a baboon, Papio cynocephalus, was determined by automated sequencing of the intact chain and six fragments generated by specific cleavage reactions. the existence of structural heterogeneity at position 75, where both valyl and isoleucyl residues were found, is suggestive of the presence of nonallelic Vγ and Iγ-chain genes in this species, and further emphasizes the extent to which the genetic basis of hemoglobin production among many higher primates is similar. Comparison of the sequences of those γ chains from Homo sapiens, Pan troglodytes, Macaca nemestrina and P. cynocephalus that have been well characterized attests to the conservative nature of γ-chain evolution among the Anthropoidea, the differences in sequence between any two of these chains ranging from none (between the Aγ and Gγ chains of P. troglodytes and H. sapiens) to no more than five (between the Vγ chains of p. cynocephalus and the Aγ chains of H. sapiens).

Hemogloblin α-gene duplication in macaques: Individual Macaca nemestrina with three structurally different α chains

Abstract The diversity of hemoglobin phenotypes observed among Malaysian Macaca nemestrina (pig-tailed macaques) has been attributed, in part, to the presence of duplicated α-chain loci in some members of this species. To date, evidence in support of this view has been indirect, consisting of variation in proportions of α I (Asp 71 , Gln 78 ) and α II (Asp 71 , His 78 ) chains among presumptive heterozygotes. However, the discovery that erythrocytes from some M. nemestrina contain α I and α II chains in company with a third, or α III , chain (Gly 71 , Gln 78 ) provides direct evidence of duplicated α-chain loci.

An improved staining technique for precipitin bands in agar or agarose gels

Abstract A method for the staining of proteins in agar and agarose gels using three stains simultaneously and a mordant is described. When compared with conventional Coomassie brilliant blue R-250 staining procedures, it requires a comparable time expenditure but has the following advantages: 1) it is threefold to fourfold more sensitive; 2) there is increased photographic resolution on conventional photographic material; and 3) the stain has a long shelf-life and does not fade under normal lighting conditions. Conditions for the washing and drying of gels are discussed.