Cloning and expression of a sialic acid-binding lectin from the snail Cepaea hortensis.
Highly degenerated gene-specific oligonucleotide primers (GSPs) were constructed from the amino acid sequence of tryptic fragments produced from the purified sialic acid-specific lectin of the garden snail Cepaea hortensis. From the albumin glands, the total RNA or the mRNA was prepared. Combination of a universal primer with the GSPs delivered gene-specific fragments of about 650, 620 and 280 bp by polymerase chain reaction (PCR). These fragments were cloned into the vector pDrive (Qiagen) and sequenced. The resulting cDNA sequence consisted of 744 bp, including an open reading frame of 480 bp. The encoded protein consists of 159 amino acids, including the putative signal sequence peptide. The mature protein should comprise 141 amino acid residues with a calculated molecular mass of 15,529 Da. The expression of the recombinant lectin in Escherichia coli resulted in a soluble protein reacting specifically with rabbit antiserum raised against the native lectin.
Gerlach
Dieter
D
Friedrich-Schiller-University of Jena, Institute of Medical Microbiology, Semmelweisstr 4, D-07743 Jena, Germany. dieter.gerlach@med.uni-jena.de
Schlott
Bernhard
B
Schmidt
Karl-Herman
KH
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D003001:Cloning, Molecular; D018076:DNA, Complementary / Q000737:chemistry / Q000302:isolation & purification; D004925:Escherichia / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D037102:Lectins / Q000737:chemistry / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D008969:Molecular Sequence Data; D008970:Molecular Weight; D016366:Open Reading Frames; D011110:Polymorphism, Genetic; D011401:Promoter Regions, Genetic; D021382:Protein Sorting Signals; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000276:immunology; D017422:Sequence Analysis, DNA; D017385:Sequence Homology; D012908:Snails
D018076:DNA, Complementary; D037102:Lectins; D021382:Protein Sorting Signals; D011994:Recombinant Proteins; C045831:agglutinin-I, cepaea hortensis