Isolectins from Solanum tuberosum with different detailed carbohydrate binding specificities: unexpected recognition of lactosylceramide by N-acetyllactosamine-binding lectins.
Glycosphingolipid recognition by two isolectins from Solanum tuberosum was compared by the chromatogram binding assay. One lectin (PL-I) was isolated from potato tubers by affinity chromatography, and identified by MALDI-TOF mass spectrometry as a homodimer with a subunit molecular mass of 63,000. The other (PL-II) was a commercial lectin, characterized as two homodimeric isolectins with subunit molecular masses of 52,000 and 55,000, respectively. Both lectins recognized N-acetyllactosamine-containing glycosphingolipids, but the fine details of their carbohydrate binding specificities differed. PL-II preferentially bound to glycosphingolipids with N-acetyllactosamine branches, as Galbeta4GlcNAcbeta6(Galbeta4GlcNAcbeta3)Galbeta4Glcbeta1C er. PL-I also recognized this glycosphingolipid, but bound equally well to the linear glycosphingolipid Galbeta4GlcNAcbeta3Galbeta4GlcNAcbeta3Galbeta4Glcbeta1Cer. Neolactotetraosylceramide and the B5 pentaglycosylceramide were also bound by PL-I, while other glycosphingolipids with only one N-acetyllactosamine unit were non-binding. Surprisingly, both lectins also bound to lactosylceramide, with an absolute requirement for sphingosine and non-hydroxy fatty acids. The inhibition of binding to both lactosylceramide and N-acetyllactosamine-containing glycosphingolipids by N-acetylchitotetraose suggests that lactosylceramide is also accomodated within the N-acetylchitotetraose/N-acetyllactosamine-binding sites of the lectins. Through docking of glycosphingolipids onto a three-dimensional model of the PL-I hevein binding domain, a Galbeta4GlcNAcbeta3Galbeta4 binding epitope was defined. Furthermore, direct involvement of the ceramide in the binding of lactosylceramide was suggested.
Ciopraga
J
J
Institute of Biochemistry, Romanian Academy, P.O. Box 78 200 Bucharest, 2, Romania.
Angström
J
J
Bergström
J
J
Larsson
T
T
Karlsson
N
N
Motas
C
C
Gozia
O
O
Teneberg
S
S
D000606:Amino Sugars / Q000378:metabolism*; D015703:Antigens, CD; D023181:Antimicrobial Cationic Peptides; D050260:Carbohydrate Metabolism; D002240:Carbohydrate Sequence; D004591:Electrophoresis, Polyacrylamide Gel; D007790:Lactosylceramides / Q000378:metabolism*; D037102:Lectins / Q000737:chemistry / Q000302:isolation & purification* / Q000378:metabolism; D009682:Magnetic Resonance Spectroscopy; D008958:Models, Molecular; D008969:Molecular Sequence Data; D037121:Plant Lectins; D010940:Plant Proteins / Q000737:chemistry; D016415:Sequence Alignment; D011198:Solanum tuberosum / Q000737:chemistry*; D013329:Structure-Activity Relationship
D000606:Amino Sugars; D015703:Antigens, CD; D023181:Antimicrobial Cationic Peptides; D007790:Lactosylceramides; D037102:Lectins; D037121:Plant Lectins; D010940:Plant Proteins; C045875:hevein; C000458:N-acetyllactosamine; C009744:CDw17 antigen