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Biochem J. Aug 15, 1988; 254(1): 195–202.
PMCID: PMC1135056

Chemical-modification studies of a unique sialic acid-binding lectin from the snail Achatina fulica. Involvement of tryptophan and histidine residues in biological activity.

Abstract

A unique sialic acid-binding lectin, achatininH (ATNH) was purified in single step from the haemolymph of the snail Achatina fulica by affinity chromatography on sheep submaxillary-gland mucin coupled to Sepharose 4B. The homogeneity was checked by alkaline gel electrophoresis, immunodiffusion and immunoelectrophoresis. Amino acid analysis showed that the lectin has a fairly high content of acidic amino acid residues (22% of the total). About 1.3% of the residues are half-cystine. The glycoprotein contains 21% carbohydrate. The unusually high content of xylose (6%) and fucose (2.7%) in this snail lectin is quite interesting. The protein was subjected to various chemical modifications in order to detect the amino acid residues and carbohydrate residues present in its binding sites. Modification of tyrosine and arginine residues did not affect the binding activity of ATNH; however, modification of tryptophan and histidine residues led to a complete loss of its biological activity. A marked decrease in the fluorescence emission was found as the tryptophan residues of ATNH were modified. The c.d. data showed the presence of an identical type of conformation in the native and modified agglutinin. The modification of lysine and carboxy residues partially diminished the biological activity. The activity was completely lost after a beta-elimination reaction, indicating that the sugars are O-glycosidically linked to the glycoprotein's protein moiety. This result confirms that the carbohydrate moiety also plays an important role in the agglutination property of this lectin.

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Selected References

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  • Allen AK, Neuberger A. The quantitation of glucosamine and galactosamine in glycoproteins after hydrolysis in p-toluenesulphonic acid. FEBS Lett. 1975 Dec 1;60(1):76–80. [PubMed]
  • Allen AK, Desai NN, Neuberger A. Purification of the glycoprotein lectin from the broad bean (Vicia faba) and a comparison of its properties with lectins of similar specificity. Biochem J. 1976 Apr 1;155(1):127–135. [PMC free article] [PubMed]
  • Andersen TT, Ebner KE. Reaction of the histidines of prolactin with ethoxyformic anhydride. A binding site modification. J Biol Chem. 1979 Nov 10;254(21):10995–10999. [PubMed]
  • Ashford D, Menon R, Allen AK, Neuberger A. Studies on the chemical modification and potato (Solanum tuberosum) lectin and its effect on haemagglutinating activity. Biochem J. 1981 Nov 1;199(2):399–408. [PMC free article] [PubMed]
  • Basu S, Sarkar M, Mandal C. A single step purification of a sialic acid binding lectin (AchatininH) from Achatina fulica snail. Mol Cell Biochem. 1986 Aug;71(2):149–157. [PubMed]
  • Bishayee S, Dorai DT. Isolation and characterisation of a sialic acid-binding lectin (carcinoscorpin) from Indian horseshoe crab Carcinoscorpius rotunda cauda. Biochim Biophys Acta. 1980 May 29;623(1):89–97. [PubMed]
  • Bredderman PJ. Tryptophan analysis of proteins in 6M guanidine hydrochloride: modification for more general application. Anal Biochem. 1974 Sep;61(1):298–301. [PubMed]
  • Carraway KL, Koshland DE., Jr Reaction of tyrosine residues in proteins with carbodiimide reagents. Biochim Biophys Acta. 1968 Jun 26;160(2):272–274. [PubMed]
  • Chambers RE, Clamp JR. An assessment of methanolysis and other factors used in the analysis of carbohydrate-containing materials. Biochem J. 1971 Dec;125(4):1009–1018. [PMC free article] [PubMed]
  • Chowdhury M, Sarkar M, Mandal C. Identification and isolation of an agglutinin from uterus of rats. Biochem Biophys Res Commun. 1985 Aug 15;130(3):1301–1307. [PubMed]
  • DAVIS BJ. DISC ELECTROPHORESIS. II. METHOD AND APPLICATION TO HUMAN SERUM PROTEINS. Ann N Y Acad Sci. 1964 Dec 28;121:404–427. [PubMed]
  • Dixon HB, Perham RN. Reversible blocking of amino groups with citraconic anhydride. Biochem J. 1968 Sep;109(2):312–314. [PMC free article] [PubMed]
  • Downs F, Herp A, Moschera J, Pigman W. Beta-elimination and reduction reactions and some applications of dimethylsulfoxide on submaxillary glycoproteins. Biochim Biophys Acta. 1973 Nov 11;328(1):182–192. [PubMed]
  • Edelhoch H. Spectroscopic determination of tryptophan and tyrosine in proteins. Biochemistry. 1967 Jul;6(7):1948–1954. [PubMed]
  • Goldstein IJ, Hayes CE. The lectins: carbohydrate-binding proteins of plants and animals. Adv Carbohydr Chem Biochem. 1978;35:127–340. [PubMed]
  • Habeeb AF. Preparation of enzymically active, water-insoluble derivatives of trypsin. Arch Biochem Biophys. 1967 Mar;119(1):264–268. [PubMed]
  • Hall JL, Rowlands DT., Jr Heterogeneity of lobster agglutinins. II. Specificity of agglutinin-erythrocyte binding. Biochemistry. 1974 Feb 12;13(4):828–832. [PubMed]
  • Hatcher VB, Schwarzmann GO, Jeanloz RW, McArthur JW. Purification, properties, and partial structure elucidation of a high-molecular-weight glycoprotein from cervical mucus of the bonnet monkey (Macaca radiata). Biochemistry. 1977 Apr 5;16(7):1518–1524. [PubMed]
  • Herrmann MS, Behnke WD. Physical studies on three lectins from the seeds of Abrus precatorius. Biochim Biophys Acta. 1980 Jan 24;621(1):43–52. [PubMed]
  • Hoare DG, Koshland DE., Jr A method for the quantitative modification and estimation of carboxylic acid groups in proteins. J Biol Chem. 1967 May 25;242(10):2447–2453. [PubMed]
  • Jordan F, Bassett E, Redwood WR. Proton magnetic resonance studies on wheat germ agglutinin-amino sugar interaction. Evidence for involvement of a tryptophan residue in the binding process. Biochem Biophys Res Commun. 1977 Apr 25;75(4):1015–1021. [PubMed]
  • Kelly RC, von Hippel PH. DNA "melting" proteins. III. Fluorescence "mapping" of the nucleic acid binding site of bacteriophage T4 gene 32-protein. J Biol Chem. 1976 Nov 25;251(22):7229–7239. [PubMed]
  • Lowe G, Whitworth AS. A kinetic and fluorimetric investigation of papain modified at tryptophan-69 and -177 by N-bromosuccinimide. Biochem J. 1974 Aug;141(2):503–515. [PMC free article] [PubMed]
  • LOWRY OH, ROSEBROUGH NJ, FARR AL, RANDALL RJ. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed]
  • Mandal C, Basu S. An unique specificity of a sialic acid binding lectin AchatininH, from the hemolymph of Achatina fulica snail. Biochem Biophys Res Commun. 1987 Oct 29;148(2):795–801. [PubMed]
  • Marchialonis JJ, Edelman GM. Isolation and characterization of a hemagglutinin from Limulus polyphemus. J Mol Biol. 1968 Mar 14;32(2):453–465. [PubMed]
  • Means GE, Feeney RE. Reductive alkylation of amino groups in proteins. Biochemistry. 1968 Jun;7(6):2192–2201. [PubMed]
  • Melchior WB, Jr, Fahrney D. Ethoxyformylation of proteins. Reaction of ethoxyformic anhydride with alpha-chymotrypsin, pepsin, and pancreatic ribonuclease at pH 4. Biochemistry. 1970 Jan 20;9(2):251–258. [PubMed]
  • Miller RL, Collawn JF, Jr, Fish WW. Purification and macromolecular properties of a sialic acid-specific lectin from the slug Limax flavus. J Biol Chem. 1982 Jul 10;257(13):7574–7580. [PubMed]
  • Mitra D, Sarkar M, Allen AK. Further characterization of the cold agglutinin from the snail Achatina fulica. Biochem J. 1987 Mar 1;242(2):331–338. [PMC free article] [PubMed]
  • Mukherji S, Bhaduri A. UDP-glucose 4-epimerase from Saccharomyces fragilis. Presence of an essential arginine residue at the substrate-binding site of the enzyme. J Biol Chem. 1986 Apr 5;261(10):4519–4524. [PubMed]
  • Patthy L, Smith EL. Identification of functional arginine residues in ribonuclease A and lysozyme. J Biol Chem. 1975 Jan 25;250(2):565–569. [PubMed]
  • Ravindranath MH, Higa HH, Cooper EL, Paulson JC. Purification and characterization of an O-acetylsialic acid-specific lectin from a marine crab Cancer antennarius. J Biol Chem. 1985 Jul 25;260(15):8850–8856. [PubMed]
  • Roche AC, Monsigny M. Limulin (Limulus polyphemus lectin). Isolation, physiocochemical properties, sugar specificity and mitogenic activity. Prog Clin Biol Res. 1979;29:603–616. [PubMed]
  • SAIFER A, GERSTENFELD S. Photometric determination of sialic acids in serum and cerebrospinal fluid with the thiobarbituric acid method. Clin Chim Acta. 1962 Jul;7:467–475. [PubMed]
  • Sarkar M, Bachhawat BK, Mandal C. A new cold agglutinin from Achatina fulica snails. Arch Biochem Biophys. 1984 Aug 15;233(1):286–289. [PubMed]
  • Spande TF, Green NM, Witkop B. The reactivity toward N-bromosuccinimide of tryptophan in enzymes, zymogens, and inhibited enzymes. Biochemistry. 1966 Jun;5(6):1926–1933. [PubMed]
  • Stockert RJ, Morell AG, Scheinberg IH. Mammalian hepatic lectin. Science. 1974 Oct 25;186(4161):365–366. [PubMed]
  • Tsai CM, Zopf DA, Yu RK, Wistar R, Jr, Ginsburg V. A Waldenström macroglobulin that is both a cold agglutinin and a cryoglobulin because it binds N-acetylneuraminosyl residues. Proc Natl Acad Sci U S A. 1977 Oct;74(10):4591–4594. [PMC free article] [PubMed]
  • van Kuik JA, van Halbeek H, Kamerling JP, Vliegenthart JF. Primary structure of the low-molecular-weight carbohydrate chains of Helix pomatia alpha-hemocyanin. Xylose as a constituent of N-linked oligosaccharides in an animal glycoprotein. J Biol Chem. 1985 Nov 15;260(26):13984–13988. [PubMed]
  • WARREN L. The thiobarbituric acid assay of sialic acids. J Biol Chem. 1959 Aug;234(8):1971–1975. [PubMed]

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