Skip to main content
NCBI home page
Biochemical Journal logoLink to Biochemical Journal
. 1978 Jul 1;173(1):321–330. doi: 10.1042/bj1730321

Primary structure of the major β-chain of rat haemoglobins

Laura M Garrick 1, Richard L Sloan 1, Thomas W Ryan 1, Thomas J Klonowski 1, Michael D Garrick 1,*
PMCID: PMC1185777  PMID: 687373

Abstract

The amino acid sequence of the major β-chain, IIβ, from rat haemoglobins was established with an automated sequencer. Amino acid heterogeneities were found that appear to result from allelic variation at particular residues. We applied several new or unusual techniques in determining the sequence: (1) reaction of the polypeptide with dansylaziridine for detection of cysteine; (2) blockage of the N-terminal residue and the ε-amino group of lysine residues with 1-fluoro-2-nitro-4-trimethylammoniobenzene iodide and subsequent identification of the modified lysine phenylthiohydantoin by absorbance at 420nm; (3) identification of histidine phenylthiohydantoin by its blue fluorescence under long-wave u.v. light; (4) cleavage of the chain into two or three fragments and subsequent sequencing without purification [a detailed statement giving the major phenylthiohydantoins assigned at each step for each sequence run before their alignment in individual sequences has been deposited as Supplementary Publication SUP 50084 (10 pages) at the British Library Lending Division, Boston Spa, Wetherby, West Yorkshire LS23 7BQ, U.K., from whom copies can be obtained on the terms indicated in Biochem. J. (1978) 169, 5]; (5) separation of fragments produced by CNBr cleavage by cation-exchange chromatography; (6) peptide sequencing after attachment of the peptide to cytochrome c. The amino acid sequence was confirmed by amino acid compositions of the complete chain, of CNBr fragments 1 and 3, and of 11 purified tryptic peptides.

Full text

PDF
321

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bricker J., Garrick M. D. An isoleucine-valine substitution in the beta chain of rabbit hemoglobin. Biochim Biophys Acta. 1974 Jun 7;351(2):437–441. doi: 10.1016/0005-2795(74)90208-6. [DOI] [PubMed] [Google Scholar]
  2. Chua C. G., Carrell R. W., Howard B. H. The amino acid sequence of the alpha chain of the major haemoglobin of the rat (Rattus norvegicus). Biochem J. 1975 Jul;149(1):259–269. doi: 10.1042/bj1490259. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Chua C. G., Carrell R. W. Three cysteine residues in the alpha chain of rat haemoglobin (albino Rattus norvegicus): 13 (A 11), 104 (G 11) and 111 (G 18). Biochim Biophys Acta. 1974 Oct 9;365(2):328–334. doi: 10.1016/0005-2795(74)90005-1. [DOI] [PubMed] [Google Scholar]
  4. Clegg J. B., Naughton M. A., Weatherball D. J. Abnormal human haemoglobins. Separation and characterization of the alpha and beta chains by chromatography, and the determination of two new variants, hb Chesapeak and hb J (Bangkok). J Mol Biol. 1966 Aug;19(1):91–108. doi: 10.1016/s0022-2836(66)80052-9. [DOI] [PubMed] [Google Scholar]
  5. Edman P., Begg G. A protein sequenator. Eur J Biochem. 1967 Mar;1(1):80–91. doi: 10.1007/978-3-662-25813-2_14. [DOI] [PubMed] [Google Scholar]
  6. Garrick L. M., Sharma V. S., McDonald M. J., Ranney H. M. Rat haemoglobin heterogeneity. Two structurally distinct alpha chains and functional behaviour of selected components. Biochem J. 1975 Jul;149(1):245–258. doi: 10.1042/bj1490245. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Garrick M. D., Hafner R., Bricker J., Garrick L. M. Genetic variation in the primary structure of the beta chain of rabbit hemoglobin. Ann N Y Acad Sci. 1974 Nov 29;241(0):436–438. doi: 10.1111/j.1749-6632.1974.tb21899.x. [DOI] [PubMed] [Google Scholar]
  8. Garrick M. D., Sloan R. L. Covalent attachment of peptides to cytochrome C for automated sequence determination. Prep Biochem. 1977;7(2):111–128. doi: 10.1080/00327487708061630. [DOI] [PubMed] [Google Scholar]
  9. Gray W. R. Protein structure: a new strategy for sequence analysis. Nature. 1968 Dec 28;220(5174):1300–1304. doi: 10.1038/2201300a0. [DOI] [PubMed] [Google Scholar]
  10. Hermodson M. A., Ericsson L. H., Titani K., Neurath H., Walsh K. A. Application of sequenator analyses to the study of proteins. Biochemistry. 1972 Nov 21;11(24):4493–4502. doi: 10.1021/bi00774a011. [DOI] [PubMed] [Google Scholar]
  11. Houmard J., Drapeau G. R. Staphylococcal protease: a proteolytic enzyme specific for glutamoyl bonds. Proc Natl Acad Sci U S A. 1972 Dec;69(12):3506–3509. doi: 10.1073/pnas.69.12.3506. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Inglis A. S., Nicholls P. W. Subnanomole detection of phenylthiohydantoins of amino acids after thin-layer chromatography. J Chromatogr. 1974 Oct 23;97(2):289–292. doi: 10.1016/s0021-9673(00)95611-5. [DOI] [PubMed] [Google Scholar]
  13. Jacobson G. R., Schaffer M. H., Stark G. R., Vanaman T. C. Specific chemical cleavage in high yield at the amino peptide bonds of cysteine and cystine residues. J Biol Chem. 1973 Oct 10;248(19):6583–6591. [PubMed] [Google Scholar]
  14. Perutz M. F., Muirhead H., Cox J. M., Goaman L. C. Three-dimensional Fourier synthesis of horse oxyhaemoglobin at 2.8 A resolution: the atomic model. Nature. 1968 Jul 13;219(5150):131–139. doi: 10.1038/219131a0. [DOI] [PubMed] [Google Scholar]
  15. SANGER F., TUPPY H. The amino-acid sequence in the phenylalanyl chain of insulin. I. The identification of lower peptides from partial hydrolysates. Biochem J. 1951 Sep;49(4):463–481. doi: 10.1042/bj0490463. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Sutton D. A., Drewes S. E., Welz U. Evaluation of 1-fluoro-2-nitro-4-trimethylammoniobenzene iodide, a protein-solubilizing reagent. Biochem J. 1972 Nov;130(2):589–595. doi: 10.1042/bj1300589. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Biochemical Journal are provided here courtesy of The Biochemical Society

RESOURCES