Skip to main content
SpringerLink
Log in

Comparative Studies of Two Araceous Lectins by Steady State and Time-Resolved Fluorescence and CD Spectroscopy

  • Original Paper
  • Published:
Journal of Fluorescence Aims and scope Submit manuscript

Abstract

Transitions in the tryptophan microenvironment and secondary structure of two monocot lectins from Sauromatum guttatum and Arisaema tortuosum under different denaturing conditions were studied by steady state and time resolved fluorescence and CD spectroscopy. The lectins exist as tetramers with a single tryptophan residue estimated per monomer, present in a polar environment. Quenching with ionic quenchers showed predominantly electropositive environment for tryptophan residues. Acrylamide had maximum quenching effect. A decrease in KI quenching due to lectin denaturation indicated redistribution of charges as a result of possible conformational change. The two values for lifetimes of tryptophanyl population (1.2–1.4 and 6.3–6.4 ns) reduced substantially on quenching or denaturation. Similarly, both the lectins showed a drastic loss of secondary structure in 5 M Gdn-HCl or 6 M Urea or at pH 2.0 and below. For the first time araceous lectins, like legume lectins are shown to bind adenine. The presence of a compact structure at alkaline pH 10.0–12.0 was observed in CD spectra.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2
Fig. 3
Fig. 4

Similar content being viewed by others

References

  1. Burstein EA, Vedenkina NS, Ivkova MN (1973) Fluorescence and the location of tryptophan residues in protein molecules. Photochem Photobiol 18(4):263–279 doi:10.1111/j.1751-1097.1973.tb06422.x

    Article  PubMed  CAS  Google Scholar 

  2. Lakowicz EM, Weber G (1973) Quenching of protein fluorescence by oxygen. Detection of structural fluctuations in proteins on the nanosecond time scale. Biochemistry 12:4171–4179 doi:10.1021/bi00745a021

    Article  PubMed  CAS  Google Scholar 

  3. Demchenko AP (1986) Fluorescence analysis of protein dynamics. Essays Biochem 22:120–157

    PubMed  CAS  Google Scholar 

  4. Wright LM, Wood SD, Reynolds CD, Rizkallah PJ, Peumans WJ, Van Damme EJM et al (1996) Purification, crystallization and preliminary X-ray analysis of a mannose-binding lectin from bluebell (Scilla campanulata) bulbs. Acta Crystallogr D52:1021–1023

    CAS  Google Scholar 

  5. Van Damme EJM, Allen AK, Peumans WJ (1988) Related mannose-specific lectins from different species of the family Amaryllidaceae. Physiol Plant 73:52–57 doi:10.1111/j.1399-3054.1988.tb09192.x

    Article  Google Scholar 

  6. Peumans WJ, Kellens JT, Allen AK, Van Damme EJM (1991) Isolation and characterization of a seed lectin from elderberry (Sambucus nigra L.) and its relationship to the bark lectins. Carbohydr Res 213:7–17 doi:10.1016/S0008-6215(00)90593-7

    Article  PubMed  CAS  Google Scholar 

  7. Van Damme EJM, Goossens K, Smeets K, Van Leuven F, Erhaert P, Peumans WJ (1995) The major tuber storage protein of araceae species is a lectin. Characterization and molecular cloning of the lectin from Arum maculatum, L. Plant Physiol 107:1147–1158 doi:10.1104/pp.107.4.1147

    Article  PubMed  Google Scholar 

  8. Van Damme EJM, Peumans WJ, Barre A, Rougé P (1998) Plant lectins: A composite of several distinct families of structurally and evolutionary related proteins with diverse biological roles. Crit Rev Plant Sci 17:645–662 doi:10.1016/S0735-2689(98)00365-7

    Google Scholar 

  9. Kamboj SS, Shangary S, Singh J, Kamboj KK, Sandhu RS (1995) New lymphocyte stimulating monocot lectins from family Araceae. Immunol Invest 24(5):845–855 doi:10.3109/08820139509060711

    Article  PubMed  CAS  Google Scholar 

  10. Shangary S, Singh J, Kamboj SS, Kamboj KK, Sandhu RS (1995) Purification and properties of four monocot lectins from the family araceae. Phytochemistry 40:449–455 doi:10.1016/0031-9422(95)00229-Z

    Article  PubMed  CAS  Google Scholar 

  11. Dhuna V, Bains JS, Kamboj SS, Singh J, Shanmugavel, Saxena AJ (2005) Purification and Characterization of a Lectin from Arisaema tortuosum Schott Having in-vitro Anticancer Activity against Human Cancer cell lines. J Biochem Mol Biol 38(5):526–532

    PubMed  CAS  Google Scholar 

  12. Kaur A, Kamboj SS, Singh J, Saxena AK, Dhuna V (2005) Isolation of a novel N-acetyl-D-lactosamine specific lectin from Alocasia cucullata (Schott.). Biotechnol Lett 27:1815–18220 doi:10.1007/s10529-005-3559-y

    Article  PubMed  CAS  Google Scholar 

  13. Singh J, Kamboj SS, Sandhu RS, Shangary S, Kamboj KK (1993) Purification and characterization of a tuber lectin from Alocasia indica. Phytochemistry 33:979–983 doi:10.1016/0031-9422(93)85007-E

    Article  CAS  Google Scholar 

  14. Allen AK, Neurberger A, Sharon N (1973) The purification, composition and specificity of wheat-germ agglutinin. Biochem J 131(1):155–162

    PubMed  CAS  Google Scholar 

  15. Kamboj SS, Shangary S, Singh J, Kamboj KK, Sandhu RS (1995) New lymphocyte stimulating monocot lectins from family Araceae. Immunol. Invest. 24(5):845–855 doi:10.3109/08820139509060711

    Article  PubMed  CAS  Google Scholar 

  16. Shangary S, Kamboj SS, Singh J, Kamboj KK, Sandhu RS (1996) New lymphocyte stimulating monocot lectins from family Araceae. Immunol. Invest. 25(4):273–278 doi:10.3109/08820139609059310

    Article  PubMed  CAS  Google Scholar 

  17. Singh J, Kamboj SS (2004) A novel mitogenic and antiproliferative lectin from a wild cobra lily, Arisaema flavum. Biochem Biophys Res Commun 318:1057–1065 doi:10.1016/j.bbrc.2004.04.135

    Article  PubMed  CAS  Google Scholar 

  18. Kaur M, Rup PJ, Saxena AK, Khan RH, Ashraf MT, Kamboj SS et al (2006) A tuber lectin from Arisaema helleborifolium Schott with anti-insect activity against melon fruit Xy, Bactrocera cucurbitae (Coquillett) and anti-cancer effect on human cancer cell lines. Arch Biochem Biophys 445:156–165 doi:10.1016/j.abb.2005.10.021

    Article  PubMed  CAS  Google Scholar 

  19. Dhuna V, Bains JS, Kamboj SS, Singh J, Shanmugavel, Saxena AJ (2007) Purification and Characterization of a Lectin from Arisaema tortuosum Schott having in-vitro Anticancer Activity against Human Cancer Cell Lines. J Biochem Mol Biol 38:526–532

    Google Scholar 

  20. Spande TF, Witkop B (1967) Determination of the tryptophan content of proteins with N-bromosuccinimide. Methods Enzymol 11:498–506 doi:10.1016/S0076-6879(67)11060-4

    Article  CAS  Google Scholar 

  21. Ali V, Prakash K, Kulkarni S, Ahmad A, Madhusudan KP, Bhakuni V (1999) 8-anilino-1-naphthalene sulfonic acid (ANS) induces folding of acid unfolded cytochrome c to molten globule state as a result of electrostatic interactions. Biochemistry 38(41):13635–13642 doi:10.1021/bi9907835

    Article  PubMed  CAS  Google Scholar 

  22. Gasymov OK, Glasgow BJ (2007) ANS fluorescence: Potential to augment the identification of the external binding sites of proteins. Biochim Biophys Acta 1774(3):403–411

    PubMed  CAS  Google Scholar 

  23. Watanabe K, Honjo E, Tsukamoto T, Funatsu G (1992) Fluorescence studies on the interaction of adenine with ricin A chain. FEBS Lett 304:249–251 doi:10.1016/0014-5793(92)80630-Y

    Article  PubMed  CAS  Google Scholar 

  24. Chipman DM, Grisaro V, Sharon N (1967) The binding of oligosaccharides containing N-acetylglucosamine and N-acetylmuramic acid to lysozyme. J Biol Chem 242:4388–4394

    PubMed  CAS  Google Scholar 

  25. Siddiqui MZ, Sharma AK, Kumar S (1996) Solution conformation of tuftsin. Int J Biol Macromol 19(2):99–102 doi:10.1016/0141-8130(96)01108-7

    Article  PubMed  CAS  Google Scholar 

  26. Eftink MR, Ghiron CA (1984) Indole fluorescence quenching studies on proteins and model systems: use of the inefficient quencher succinimide. Biochemistry 23:3891–3889 doi:10.1021/bi00312a016

    Article  CAS  Google Scholar 

  27. Katre UV, Suresh CG, Khan MI, Gaikwad SM (2007) Steady state and time-resolved fluorescence studies of a hemagglutinin from Moringa oleifera. J Fluoresc 18(2):479–485 doi:10.1007/s10895-007-0289-7

    Article  PubMed  Google Scholar 

  28. Komath SS, Swamy MJ (1999) Fluorescence quenching, time resolved fluorescence and chemical modification studies on the tryptophan residues of snake gourd (Trichosanthes anguina) seed lectin. J Photochem Photobiol B Biol 50:108–118 doi:10.1016/S1011-1344(99)00079-2

    Article  CAS  Google Scholar 

  29. Kenoth R, Swamy MJ (2003) Steady-state and time-resolved fluorescence studies on Trichosanthes cucumerina seed lectin. J Photochem Photobiol B Biol 69:193–201

    Article  CAS  Google Scholar 

  30. Sultan NAM, Swamy MJ (2005) Fluorescence quenching and time-resolved fluorescence studies on Trichosanthes dioica seed lectin. J Photochem Photobiol B Biol 80:93–100 doi:10.1016/j.jphotobiol.2005.03.003

    Article  CAS  Google Scholar 

  31. Sultan NAM, Rao RN, Nadimpalli SK, Swamy MJ (2006) Tryptophan environment, secondary structure and thermal unfolding of the galactose-specific seed lectin from Dolichos lablab: Fluorescence and circular dichroism spectroscopic studies. Biochim Biophys Acta 1760:1001–1008

    PubMed  CAS  Google Scholar 

  32. Lehrer SS (1971) Solute perturbation of protein fluorescence. The quenching of tryptophyl fluorescence of model compounds and of lysozyme by iodide ion. Biochemistry 10:3254–3263 doi:10.1021/bi00793a015

    Article  PubMed  CAS  Google Scholar 

  33. Etzler ME, Gupta S, Borrebaeck C (1981) Carbohydrate binding properties of the Dolichos biflorus lectin and its subunits. J Biol Chem 256:2367–2370

    PubMed  CAS  Google Scholar 

  34. Gegg CV, Roberts DD, Segel IH, Etzler ME (1992) Characterization of the adenine binding sites of two Dolichos biflorus Lectins. Biochemistry 31:6938–6942 doi:10.1021/bi00145a011

    Article  PubMed  CAS  Google Scholar 

  35. Roberts DD, Goldstein IJ (1983) Adenine binding sites of the lectin from lima beans (Phaseolus lunatus). J Biol Chem 258:13820–13824

    PubMed  CAS  Google Scholar 

  36. Puri KD, Surolia A (1994) Amino acid sequence of the Winged Bean (Psophocarpus tetragonolobus) basic lectin. Adenine binding and identification of the active-site tryptophan residue. J Biol Chem 269:30917–30926

    PubMed  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Biochemistry, National Chemical Laboratory, Pune, 411008, Maharashtra, India

    Poorva N. Dharker, Sushama M. Gaikwad, C. G. Suresh & M. Islam Khan

  2. Department of Molecular Biology and Biochemistry, Guru Nanak Dev University, Amritsar, 143005, Punjab, India

    Vikram Dhuna, Jatinder Singh & Sukhdev Singh Kamboj

Authors
  1. Poorva N. Dharker
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Sushama M. Gaikwad
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. C. G. Suresh
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Vikram Dhuna
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. M. Islam Khan
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Jatinder Singh
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Sukhdev Singh Kamboj
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Sushama M. Gaikwad.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Dharker, P.N., Gaikwad, S.M., Suresh, C.G. et al. Comparative Studies of Two Araceous Lectins by Steady State and Time-Resolved Fluorescence and CD Spectroscopy. J Fluoresc 19, 239–248 (2009). https://doi.org/10.1007/s10895-008-0409-z

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s10895-008-0409-z

Keywords

Search

Navigation