Characterization of serum immunoglobulin M of grouper and cDNA cloning of its heavy chain.
Immunoglobulin M (IgM) from the whole serum of grouper fish, Epinephelus coioides was purified by affinity chromatography using protein A-Sepharose column. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) under reducing conditions revealed that the relative molecular masses (Mr) of the equimolar heavy and light chains of IgM were 78,000 and 27,000, respectively. The cDNAs encoding IgM heavy chain comprising its variable (VH) and constant (CH) regions have been cloned and sequenced from a grouper kidney cDNA library by antibody screening method. Five VH (130-142 amino acids) and four CH (450-454 amino acids) families were identified. The variable and constant regions were conserved with their putative domains. All the four constant region domains (CH1-CH2-CH3-CH4) contained each three conserved cysteine residues, which are considered to form the inter- and intra-chain disulfide bridges. There were three carbohydrate acceptor sites in the constant region. In general, the pattern of IgM gene organization seems to resemble that of other teleosts. Moreover, the CH genes in grouper IgM occur as multifamily as reported in Atlantic salmon and common carp.
Cheng
Chao-An
CA
Department of Food Science, National Kinmen Institute of Technology, Kinmen, Taiwan.
John
Joseph Abraham Christopher
JA
Wu
Ming-Shan
MS
Lee
Chiou-Yueh
CY
Lin
Chih-Hung
CH
Lin
Cheng-Hui
CH
Chang
Chi-Yao
CY
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002846:Chromatography, Affinity / Q000662:veterinary; D016000:Cluster Analysis; D007127:Immunoglobulin Constant Regions / Q000737:chemistry / Q000235:genetics; D007143:Immunoglobulin Heavy Chains / Q000737:chemistry / Q000235:genetics*; D007075:Immunoglobulin M / Q000097:blood / Q000235:genetics* / Q000276:immunology / Q000302:isolation & purification; D007135:Immunoglobulin Variable Region / Q000737:chemistry / Q000235:genetics; D008969:Molecular Sequence Data; D010473:Perciformes / Q000235:genetics* / Q000276:immunology*; D010802:Phylogeny; D012313:RNA / Q000737:chemistry / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction / Q000662:veterinary; D016415:Sequence Alignment; D017422:Sequence Analysis, DNA; D020539:Sequence Analysis, Protein
D007127:Immunoglobulin Constant Regions; D007143:Immunoglobulin Heavy Chains; D007075:Immunoglobulin M; D007135:Immunoglobulin Variable Region; D012313:RNA