A mannose-specific lectin from Vicia villosa seeds.
A lectin specific to mannose has been purified from Vicia villosa seed by (NH4)2SO4 fractionation, GalNAc-Sepharose and Man-Sepharose affinity chromatography. It was defined as VVLM, which showed a single band on an acidic-PAGE stained with Coomassie brilliant blue. The molecular weight of VVLM was 50 kDa as determined by gel filtration on Biogel P-100 column. The VVLM molecule consists of 2 distinct subunits with apparent molecular weight of 30 kDa and 22 kDa determined by SDS-PAGE. VVLM has at least four isolectins with similar haemagglutinating activity. Its extinction coefficient is calculated as A1(1cm) = 16.4 at 280 nm. Sugars could not be detected by phenol-sulfuric acid method. The circular dichroism analysis at far UV indicated that VVLM was a beta-sheet-rich protein, and gave no alpha-helix, 69% beta-sheet, 14% beta-turn by Provencher and Glockner method. The lectin was inhibited by alpha-methyl-D-mannose at 12.5 mM and glucose or GlcNAc at 50 mM. The carbohydrate binding specificity of VVLM was investigated by using affinity chromatography on a VVLM-Sepharose column. Among various Asn-linked oligosaccharides, core structure Man alpha 1-->3(Man alpha 1-->6)Man beta 1-->4GlcNAc beta 1-->4GlcNAcOT were found to have high affinity for VVLM-Sepharose. The antisera of VVLM did not produce precipitin line with VVLG in agar double diffusion plate indicating no serological relationship between VVLM and VVLG. However VVLM showed similar immunodeterminants of some other lectins of mannose specificity such as Con A, PSL, LCA and VFL.
Qian
R
R
Shanghai Institute of Biochemistry, Academia Sinica, China.
Shi
W X
WX
Shen
Z M
ZM
Sun
C
C
D000596:Amino Acids / Q000032:analysis; D002240:Carbohydrate Sequence; D002942:Circular Dichroism; D037102:Lectins / Q000737:chemistry / Q000302:isolation & purification*; D008358:Mannose / Q000737:chemistry*; D008969:Molecular Sequence Data; D015394:Molecular Structure; D008970:Molecular Weight; D009844:Oligosaccharides / Q000737:chemistry; D037121:Plant Lectins; D012639:Seeds / Q000737:chemistry*
D000596:Amino Acids; D037102:Lectins; D009844:Oligosaccharides; D037121:Plant Lectins; D008358:Mannose