Purification, characterization and cloning of a ricin B-like lectin from mushroom Clitocybe nebularis with antiproliferative activity against human leukemic T cells.
BACKGROUND
Lectins are a diverse group of carbohydrate-binding proteins exhibiting numerous biological activities and functions.


METHODS
Two-step serial carbohydrate affinity chromatography was used to isolate a lectin from the edible mushroom clouded agaric (Clitocybe nebularis). It was characterized biochemically, its gene and cDNA cloned and the deduced amino acid sequence analyzed. Its activity was tested by hemagglutination assay and carbohydrate-binding specificity determined by glycan microarray analysis. Its effect on proliferation of several human cell lines was determined by MTS assay.


RESULTS
A homodimeric lectin with 15.9-kDa subunits agglutinates human group A, followed by B, O, and bovine erythrocytes. Hemagglutination was inhibited by glycoprotein asialofetuin and lactose. Glycan microarray analysis revealed that the lectin recognizes human blood group A determinant GalNAcalpha1-3(Fucalpha1-2)Galbeta-containing carbohydrates, and GalNAcbeta1-4GlcNAc (N,N'-diacetyllactosediamine). The lectin exerts antiproliferative activity specific to human leukemic T cells.


CONCLUSIONS
The protein belongs to the ricin B-like lectin superfamily, and has been designated as C. nebularis lectin (CNL). Its antiproliferative effect appears to be elicited by binding to carbohydrate receptors on human leukemic T cells.


CONCLUSIONS
CNL is one of the few mushroom ricin B-like lectins that have been identified and the only one so far shown to possess immunomodulatory properties.
Pohleven
Jure
J
Department of Biotechnology, Jozef Stefan Institute, Jamova 39, SI-1000 Ljubljana, Slovenia. jure.pohleven@ijs.si
Obermajer
Natasa
N
Sabotic
Jerica
J
Anzlovar
Sabina
S
Sepcić
Kristina
K
Kos
Janko
J
Kralj
Bogdan
B
Strukelj
Borut
B
Brzin
Joze
J
D000363:Agaricales / Q000737:chemistry*; D000595:Amino Acid Sequence; D002240:Carbohydrate Sequence; D045744:Cell Line, Tumor; D049109:Cell Proliferation / Q000187:drug effects*; D002850:Chromatography, Gel; D018076:DNA, Complementary; D004591:Electrophoresis, Polyacrylamide Gel; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007526:Isoelectric Point; D037102:Lectins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000494:pharmacology; D015458:Leukemia, T-Cell / Q000473:pathology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D017386:Sequence Homology, Amino Acid; D021241:Spectrometry, Mass, Electrospray Ionization
D018076:DNA, Complementary; D037102:Lectins