Amphioxus P23 Gene: Cloning, Characterization, Expression, And Purification Of Recombinant Protein And Antibody Production

The molecular chaperone p23 is a cofactor of the Hsp90 protein that plays a critical role in the folding of a series of important regulators of cellular function. It is required for the proper folding of many signaling proteins and only found in eukaryotes. Amphioxus, a cephalochordate, is the closest living relative to vertebrate, and has been widely known as the most important animal to analyze the origin and evolution of vertebrates. Study on the gene structure, function and expression in amphioxus will contribute to the understanding of the origin and evolution of the vertebrates.In this paper, AmphiP23, encoding an amphioxus p23 protein, was identified from the gut cDNA library of Branchiostoma belcheri tsingtaunese. It contains a 513 bp open reading frame corresponding to a deduced protein of 170 amino acids. Phylogenetic analysis shows that vertebrate and invertebrate p23/p23-like proteins are each grouped together, with AmphiP23 falling at the base of vertebrate p23/p23-like clade, suggesting that the divergence of vertebrate and invertebrate p23 genes probably occurs prior to the split of invertebrate/vertebrate from a common ancestor around 550 million years ago. Northern blotting reveals a ubiquitous expression pattern of AmphiP23 in all tissues examined of adults, while whole mount in situ hybridization demonstrates a tissue- and stage-specific expression pattern of AmphiP23 in developing embryos and larvae. Presumably, the ubiquitous expression pattern of AmphiP23 in adult amphioxus represents the ancestral type of p23 gene prior to its split to human paralogs p23 and tsp23, while the tissue- and stage-specific expression pattern during early embryonic development implicates a role of AmphiP23 in anterior/posterior patterning.The recombinant p23 of amphioxus has been produced and purified. The cDNA...