Simple affinity chromatographic procedure to purify beta-galactoside binding lectins.
Affinity chromatography based on the commercial resin Sepharose CL-6B was used to isolate new C1-beta-type lectins from crude preparations of snake venoms (Bothrops jararaca, Bothrops jararacussu, Bothrops newiedi, Bothrops moojeni, Lachesis muta rhombeata). Most of the C-type lectins could be eluted with almost 100% recovery using the competitor isopropyl-beta-D-thiogalactoside (IPTG) or through Ca2+ sequestration with EDTA. The lectin yield varied considerably among the different snake species, but B. newiedi venom was a particularly rich source of lectin, retaining 2.7 mg of lectin by milliliter of resin in saturating conditions. C1-alpha-lectins from Crotalus durisus terrificus venom, from the jack fruit (jacalin) and from bread fruit seeds extract (frutalin) had no affinity, either with or without Ca2+ added, for Sepharose CL-6B, showing that the resin is specific for C1-beta-type lectins. Sepharose CL-6B used as galactose-affinity chromatography provides a simple and fast method for isolating C-type beta-galactoside binding lectins from crude sample preparations.
De-Simone
S G
SG
Laboratório de Bioquímica de Proteínas e Peptídeos, Departamento de Bioquímica e Biologia Molecular, Instituto Oswaldo Cruz, Fiocruz, Av Brasil 4365, 21045-900 RJ, Brazil. dsimone@ioc.fiocruz.br
Netto
C C
CC
Silva
F P
FP
D000595:Amino Acid Sequence; D000818:Animals; D017837:Bothrops / Q000378:metabolism; D002846:Chromatography, Affinity / Q000379:methods*; D003435:Crotalid Venoms / Q000032:analysis / Q000737:chemistry / Q000494:pharmacology; D037161:Galectins / Q000032:analysis* / Q000737:chemistry / Q000302:isolation & purification; D006386:Hemagglutination Tests; D006801:Humans; D012685:Sepharose / Q000031:analogs & derivatives / Q000737:chemistry; D020539:Sequence Analysis, Protein; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D014757:Viper Venoms / Q000032:analysis / Q000737:chemistry / Q000494:pharmacology; D017819:Viperidae / Q000378:metabolism
D003435:Crotalid Venoms; D037161:Galectins; D014757:Viper Venoms; C047264:sepharose CL 6B; D012685:Sepharose