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Molecular and kinetic properties of sperm specific LDH after radiation inactivation

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Abstract

Radiation inactivation of sperm specific lactate dehydrogenase-C4 (LDH-C4) has been studied and compared with the somatic LDH in aqueous solution. D37 of C isozyme was 470 Gy and that of B isozyme was 520 Gy. Semi-log plots of log N/No versus dose suggested that the inactivation of two LDH isozymes in presence of normal saline follows a single hit kinetics. Target molecular weight calculated by radiation analysis was found as 1.52 × 105 gm/mole for LDH-C4 and 1.38 × 105 gm/mole for LDH-B4. SDS-PAGE of irradiated enzymes showed a band of 35 kDa but did not indicate the presence of any other extra band, when compared with sham-irradiated enzymes. Chemical kinetics of residual activity following irradiation at D37 showed decrease in Vmax with coenzymes and primary substrates. However, decrease in Km was seen with pyruvate as increasing substrate. Nevertheless, Km did not change when NAD+ was the leading substrate for LDH-B4 or LDH-C4. A hyperchromicity in intrinsic fluorescence and a blue shift in λmax over sham-irradiated LDH-C4 revealed the exposure of buried tryptophan residues to the surface after radiation inactivation. Results suggest that inspite of presence of variant amino acids, the conformations of two isozymes are stabilized by similar forces which behave in a similar way for radiation inactivation in aqueous phase.

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References

  1. Goldberg E: Sperm specific lactate dehydrogenase and development of contraceptive vaccine. In: D.A. Clark, B.A. Croy (eds). Reproductive Immunology, Elsevier Science Publishers, Amsterdam, 1986, pp 137-140

    Google Scholar 

  2. Gupta GS, Kinsky RG: Modulation of alloimmune responses in vivo and in vitro by sperm specific lactate dehydrogenase-C4. Mol Cell Biochem 125: 145-151, 1993

    Google Scholar 

  3. Gupta GS, Chaturvedi G, Joshi A: Sex dependent immune responses by allogenic LDH isozymes. Mol Cell Biochem 152: 115-119, 1996

    Google Scholar 

  4. Gupta GS, Syal N: Immune responses of chemically modified homologous LDH-C4 and their effect on fertility regulation in mice. Am J Reprod Immunol 37: 206-211, 1997

    Google Scholar 

  5. Adelstein SJ: Coenzyme binding by X-irradiated Glutamate and Lactate Dehydrogenase. Biochem 4: 891-896, 1965

    Google Scholar 

  6. Buchanan JD, Armstrong DA: Free radical inactivation of lactate dehydrogenase. Int J Radiat Biol 30: 115-127, 1976

    Google Scholar 

  7. Saito M: Coenzyme protection of lactic dehydrogenase against inactivation by gamma rays. Int J Radiat Biol 34: 95-99, 1978

    Google Scholar 

  8. Schuessler H, Schilling K: Oxygen effect in the radiolysis of proteins. Part 2 Bovine serum albumin. Int J Radiat Biol 45: 267-281, 1984

    Google Scholar 

  9. Garrison WM: Reaction mechanisms in the radiolysis of peptides, polypeptides and proteins. Chem Rev 87: 381-398, 1987

    Google Scholar 

  10. Lee CY, Yuan JH, Goldberg E: Lactate dehydrogenase isozymes from mouse. Meth Enzymol 89D: 351-358, 1982

    Google Scholar 

  11. Gupta BL, Bhat RM: FBX dosimeter, calibration of high-dose radiation facilities (Handbook) Bhabha Atomic Research Centre, Bombay, p. 69, 1986

    Google Scholar 

  12. Hutchinson F, Pollard E: Physical principles of radiation action. In: M. Errera, A. Forssberg (eds). Mechanisms in Radiobiology Vol. 1, Academic Press, New York, 1961, pp 1-92

    Google Scholar 

  13. Dertinger H, Jung H: In: Molecular Radiation Biology. The English University Press Ltd., London, 1970

    Google Scholar 

  14. Laemilli UK: Cleavage of structural protein during the assembly of the head of bacteriophage T4. Nature 227: 680-685, 1970

    Google Scholar 

  15. Gupta GS, Kapur S, Kinsky RG: Inhibition kinetics of lactate dehydrogenase isozymes by gossypol acetic acid. Biochem Intern 17: 25-34, 1988

    Google Scholar 

  16. Cupta GS, Kang BPS: Quenching of intrinsic fluorescence of sperm specific LDH by optical isomers of gossypol. Gen Physiol Biophys 16: 215-225, 1997

    Google Scholar 

  17. Davis KA, Hatefi Y, Crawford, IP Baltscheffsky H: Spin trapping and its applications in lipid peroxidation and free radical production with liver microsomes. Arch Biochem Biophys 180: 459-464, 1977

    Google Scholar 

  18. Dranell DW, Klotz IM: Subunit constitution of proteins: A Table. Arch Biochem Biophys 166: 651-682, 1975

    Google Scholar 

  19. Nielsen M, Braestrup C: The apparent target size of rat brain benzodiazepine receptor, acetylcholineestrase and pyruvate kinase is highly influenced by experimental conditions. J Biol Chem 263: 11900-11906, 1988

    Google Scholar 

  20. Liang ZG, Shelton JA, Goldberg E: Non-cross reactivity of antibodies to murine LDH-C4 with LDH-A4 and LDH-B4. J Exp Zool 240: 377-384, 1989

    Google Scholar 

  21. Li SSL, Feldmann J, Okabe M, Pan Y CE: Molecular features and immunological properties of lactate dehydrogenase C4 isozymes from mouse and rat testes. J Bio Chem 258: 7017-7028, 1983

    Google Scholar 

  22. Pan YCE, Sharief FS, Okabe M, Huang S, Li SSL: Amino acid sequence studies on lactate dehydrogenase-C4 isozymes from mouse and rat testes. J Biol Chem 258: 7005-7016, 1983

    Google Scholar 

  23. Sakai I, Sharief FS, Steven SLL: Molecular cloning and nucleotide sequence of the cDNA for sperm specific LDH-C4 from mouse. Biochem J 242: 619-622, 1987

    Google Scholar 

  24. Adams MJ, Buchner M, Chandrashekhar K, Ford FC, Hackert MI, Li1jas A, Rossmann MG, Smiley IE, Allison WS, Everse J, Kaplan NO, Taylor SS: Structure function relationships in lactate dehydrogenase (amino acid sequence/crystallographic structure). Proc Natl Acad Sci (US) 70: 1968-1972, 1973

    Google Scholar 

  25. Winstead JA, Reece TC: Effects of gamma radiation on the chemical and physical properties of lactate dehydrogenase. Radiat Res 41: 125-134, 1970

    Google Scholar 

  26. Gupta GS, Sharma PK: Molecular inactivation of testicular hyaluronidase in solid state after proton irradiation: A study based on target size, substrate binding and thermodynamic analysis of heat denaturation. Ind J Biochem Blophys 32: 266-271, 1995

    Google Scholar 

  27. Sharma PK, Gupta GS: Catalytic properties of testicular hyaluronidase after gamma-irradiation. Radiat Environ Biophys 25: 123-133, 1986

    Google Scholar 

  28. Wright-LL, Swofford JH: Mouse lactate dehydrogenase (LDH)-C4 (Testes) is immuno-chemically cross-reactive with LDH-A4 (Muscle) and LDH-B4 (Heart). Scand J Immun 19: 247-254, 1984

    Google Scholar 

  29. Hundal MK, Syal N, Gupta GS: Immune responses of chemically modified LDH-C4. Am J Reprod Immunol 40: 259, 1998 (abstract)

    Google Scholar 

  30. O'Hern PA, Bambra CS, Isahakia M, Goldberg E: Reversible contraception in female baboon immunized with synthetic epitope of sperm specific lactate dehydrogenase. Biol Reprod 52: 331-339, 1995

    Google Scholar 

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Authors and Affiliations

  1. Department of Biophysics, Panjab University, Chandigarh, 160014, India

    G.S. Gupta & B.P.S. Kang

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  1. G.S. Gupta
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  2. B.P.S. Kang
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Gupta, G., Kang, B. Molecular and kinetic properties of sperm specific LDH after radiation inactivation. Mol Cell Biochem 206, 27–32 (2000). https://doi.org/10.1023/A:1007037128143

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