Abstract
The epsilon-NH2 groups of ovine luteinizing hormone has been modified with the long chain N-succinimidyl-3-(2-pyridyl dithiopropionate (LC-SPDP). The LC-SPDP modification primarily occurs in-NH2 groups of the α-subunit. Although, the sequential modification of lysine residue in α-subunit led to progressive reduction in the receptor binding and immunological properties but the steroidogenic activity was relatively unaffected. The immunoreactivity and receptor binding properties of LC-SPDP modified oLH molecule were more affected comparative to SPDP modified derivatives. This suggested that the increase in hydrophobic carbon chain in LC-SPDP-oLH molecules resulted into the drastic inhibition in the immunological and biological properties. However, the steroidogenic potential of LC-SPDP/or SPDP-oLH derivative was comparable. The present study clearly demonstrate that a single-NH2 group modification with LC-SPDP would generate the site for the conjugation to the toxin/carrier proteins and the resultant oLH-S-S-toxin conjugate would retain significant immunological and biological properties of the hormone molecule. (Mol Cell Biochem130: 83–90, 1994)
Similar content being viewed by others
References
Pierce JG, Parson TF: Glycoprotein hormones: structure and function. Ann Rev Biochem 50: 465–495, 1981
Rayan RJ, Charlesworth MC, McCormick DJ, Milius RP, Keutmann HT: The glycoprotein hormones: recent studies of structure function relationship. FASEB J 2: 2661–2669, 1988
Strickland TW, Puett D: Contribution of subunits to the functions of luteinizing hormone/human chorionic gonadotropin recombinants. Endocrinology 109: 1933–1942, 1981
Ward DN: Chemical approach to the structure-function relationship of the luteinizing hormone (Lutropin). In: Structure and function of the gonadotropins, McKerns KW (ed.). Plenum Publishing Corporation, pp 31–45, 1978
Sairam MR: Gonadotropic hormones, relationship between structure and function with emphasis on antagonists. In: Hormonal proteins and peptides, Li C-H (ed.). Academic Press, New York, pp 1–79, 1983
Strickland TW, Noland TD, Puett D: Gonadotropin testicular receptor interaction and subunit-subunit interactions. In: Functional correlates of hormone receptors in reproduction, Mahesh VB (ed.), Elsevier/North-Holland, Amsterdam, pp 463–479, 1980
Talwar GP, Singh O, Singh V: Birth control vaccine. In: Fertility regulation today and tomorrow, Diczfalusy E, Bygdmen E (eds). Raven Press, New York, pp 43–54, 1987
Talwar GP, Singh O, Jayashankar R, Shaha C, Suri A, Rao LV, Gaur A, Alam A, Upadhayay SN, Pal R, Chaudhuri M: Immunology S2: 93–97, 1989
Stevens VC, Cinader B, Powell JE, Lee AC, Koh SW: Preparation and formation of a antifertility vaccine, selection of peptide immunogen. Am J Reprod Immunol Microbiol 6: 307–314, 1981
Talwar GP, Sharma NC, Dubey SK, Salahuddin M, Das C, Ramakrishnan S, Hingorani V: Isoimmunization against human chorionic gonadotropin with conjugates of processed β-subunit of the hormone and tetanus toxoid. Proc Natl Acad Sci USA 73: 213–222, 1976
Talwar GP, Gupta SK, Singh O, Singh V, Das C: New approach for contraceptive vaccine. In: Advances in immunopharmacology, Vol 2, Hadden JW, Chedid L, Spreafico F, Willinghby D (eds). Pergman Press, New York, pp 415–420, 1983
Sharma NC, Singh O, Gaur A, Rao DN, Singh V, Rao LV, Seghal S, Das C, Talwar GP: Improved immunogenic formulations for antigonadotropin response. In: Immunological approaches for contraception and fertility promotion, Talwar GP (ed.). Plenum Press, New York, pp 37–41, 1986
Talwar GP, Singh O, Singh V, Rao DN, Sharma NC, Das C, Rao LV: Enhancement of anti-gonadotropin response to beta-subunit of ovine luteinizing hormone by carrier conjugation and combination with beta subunit of human chorionic gonadotropin. Fertility and Sterility 45: 120–127, 1986
Singh V: Design and synthesis of bioeffective hormonotoxins for selective elimination of gonadal cells. In: Horizons in Endocrinology, Vol III, Maggi M, Geenen V (eds). Serono Symposia series, Vol 76, Raven Press, New York, pp 197–202, 1991
Singh V: Hormonotoxin: Synthesis, characterization and bioefficacy of some defined disulfide linked conjugates of ovine lureinizing hormone with a ribosome inactivating protein, gelonin. Indian J Exp Biol 29: 916–925, 1991
Singh V, Das C:In vitro selective killing of gonadal tumor cells by hormonotoxins composed of ovine luteinizing hormone linked by a disulfide bond to a ribosome-inactivating protein, gelonin. Biochemistry International 24: 689–699, 1991
Singh V, Sairam MR: Effect of thiolation of amino groups of ovine lutropin on immunoreactivity, receptor binding and bioactivity. Mol Cell Endocrinol 63: 255–262, 1989
Sairam MR: Role of arginine residue in ovine lutropin: reversible modification by 1,2-cyclohexanedione. Arch Biochem Biophys 176: 197–203, 1976
Singh V, Sairam MR, Bhargavi GN, Akhras RG: Hormonotoxins: preparation and characterization of ovine luteinizing hormonegelonin conjugates. J Biol Chem 264: 3089–3095, 1989
Carlsson J, Drevin M, Axen R: Protein thiolation and reversible protein-protein conjugation. N-succinimidyl 3-(2-pyridyl dithio) propionate, a new heterobifunctional reagent. Biochem J 173: 723–737, 1978
Fraker PJ, Speck JC: Protein and cell membrane iodinations with a sparingly soluble chloramide 1,3,4,6-tetrachloro 3α,6α-diphenylglycouril. Biochem Biophys Res Commun 80: 849–857, 1978
Singh V: Structural requirement for the recognition of luteinizing hormone releasing hormone (LHRH) by monoclonal and conventional anti-LHRH antibodies. Biochem Cell Biol 64: 1372–1377, 1986
Singh V: Specificity and affinity characteristics of lureinizing hormone releasing hormone (LHRH) antibodies. Indian J Exp Biol 23: 673–675, 1985
Singh V: Immunospecificity and affinity studies on the monoclonal anti-gonadotropin releasing hormone (GnRH) antibodies: monoclonal produced by azotized GnRH preferentially recognise to native GnRH. Biochemistry International 15: 153–162, 1987
Singh V, Sairam MR: Hormonotoxins: conjugation of chorionic gonadotropin with ribosome-inactivating protein, gelonin and comparison with lutropin conjugate. Mol Cell Endocrinol 67: 217–229, 1989
Singh V, Sairam MR: Hormonotoxin: I. Strategy for synthesis of ovine luteinizing hormone-gelonin conjugate bearing toxin in the beta subunit. Int J Peptide Protein Res 33: 22–28, 1989
Singh V, Sairam MR: Hormonotoxins: Effects of modifying the gonadotropin alpha subunit on the generation of lutropin-toxin conjugate. Int J Peptide Protein Res 35: 46–51, 1990
Rebois RV: Establishment of gonadotropin responsive murine leydig tumor cell line. J Cell Biol 94: 70–76, 1982
Sairam MR, Li C-H: Reaction of ovine interstitial cell stimulating hormone with citraconic and maleic anhydrides. Arch Biochem Biophys 167: 534–539, 1975
Liu WK, Young KP, Nakagama, Ward DN: The role of amino group in the subunit association and receptor site interaction for ovine luteinizing hormone as studied by acylation. J Biol Chem 249: 5544–5550, 1974
Liu WK, Furlong NB, Ward DN: Effect of β-subunit acylation on lutropin receptor binding. J Biol Chem 252: 522–527, 1977
Liu WK, Esfahani M, Ward DN: Guanidination of ovine luteinizing hormone and effect on activity. Endocrine Res Commun 2: 47–63, 1975
Liu WK, Ward DN: Preparation of lutropin with acetyl or acetimidinyl substituents on the amino groups of β-subunit. Biochem Biophys Acta 405: 522–526, 1975
de la Llosa P, Durosay M, Tertrui-Clary C, Jatisz M: Chemical modification of lysine residues on ovine luteinizing hormone: Effect on biological activity. Biochem Biophys Acta 342: 97–104, 1974
Mendelson C, Dufau M, Catt K: Gonadotropin binding and stimulation of cyclic adenosine 3-5-monophosphate and testosterone production in isolated leydig cells. J Biol Chem 250: 8818–8823, 1975
Author information
Authors and Affiliations
Rights and permissions
About this article
Cite this article
Singh, V., Curtiss, R. Effect of amino group modification of ovine luteinizing hormone (oLH) by N-succinimidyl 6-[3-(2-pyridyldithio)propionate]hexanoate, a long chain N-succinimidyl-3-(2-pyridyldithio) propionate (SPDP) on immunological and biological properties: a comparative study with SPDP modified oLH. Mol Cell Biochem 130, 83–90 (1994). https://doi.org/10.1007/BF01084271
Received:
Accepted:
Issue Date:
DOI: https://doi.org/10.1007/BF01084271