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Cloning and characterization of a cDNA representing a putative complement-regulatory plasma protein from barred sand
A Dahmen1, T Kaidoh, P F Zipfel
1Department of Molecular Immunology, Bernhard Nocht Institute for Tropical Medicine, Hamburg, Germany.
Insights
Researchers identified a novel complement-regulatory protein (SBP1) in barred sand bass plasma. This protein shares structural similarities with mammalian complement proteins, offering insights into immune system evolution.
Area of Science:
- Immunology
- Evolutionary Biology
- Biochemistry
Background:
- Plasma from various vertebrates, including barred sand bass, contains molecules that cleave human complement components C3b and C4b.
- This activity is attributed to a specific protease and a cofactor protein, with the cofactor activity linked to a 110 kDa polypeptide chain of a 360 kDa plasma protein.
Purpose of the Study:
- To investigate the evolutionary conservation of complement-regulatory molecules.
- To isolate and characterize the cDNA encoding the complement-regulatory protein from barred sand bass.
Main Methods:
- Isolation of a cDNA clone (SB1) from a barred sand bass liver expression library using immunoscreening.
- Sequencing of the cDNA to determine its size and open reading frame.
- Bioinformatic analysis to predict protein structure and homology to mammalian complement-regulatory proteins.
Main Results:
- A cDNA clone (SB1) of 3397 bp was isolated, encoding a secreted protein (SBP1) of 1053 amino acid residues with a calculated mass of 115.2 kDa.
- SBP1 exhibits structural similarity to mammalian complement-regulatory proteins, organized into 17 short consensus repeats (SCRs).
- Specific SCRs of SBP1 showed homology to human factor H and human C4b-binding protein, indicating conserved functional domains.
Conclusions:
- A complete cDNA for a putative complement-regulatory protein (SBP1) has been isolated from a bony fish for the first time.
- SBP1's structural homology to mammalian proteins suggests conserved mechanisms in complement regulation across vertebrate evolution.
- This finding provides a basis for further research into fish complement systems and their evolutionary relationships with mammalian counterparts.
Abstract:
It has been demonstrated previously that plasma from a number of vertebrate species including the phylogenetically old barred sand bass possesses molecules that cleave the alpha'-chain of the activated third (C3b) and fourth (C4b) components of the human complement system. A specific protease and a cofactor protein were identified to be responsible for this cleavage. The cofactor activity in sand bass correlated with a 110 kDa polypeptide chain of a 360 kDa plasma protein. The evolutionary conservation was probed at the cDNA level and subsequently a cDNA clone of barred sand bass was isolated that represents a protein with structural similarity to mammalian complement-regulatory proteins. The cDNA (SB1) was identified by immunoscreening of a sand bass liver expression library using affinity-purified IgG antibodies raised against the isolated 110 kDa material. The cDNA is 3397 bp in size and the open reading frame represents a protein of 1053 amino acid residues with a hydrophobic signal peptide indicative of a secreted protein. The calculated mass of the mature protein (SBP1) is 115.2 kDa which is in good agreement with the molecular mass of 110 kDa determined for the sand bass serum protein. Similarly to mammalian complement-regulatory proteins, the protein deduced from the sand bass cDNA is organized into short consensus repeats (SCR). It consists of 17 SCRs, of which SCRs 2, 12 and 16 exhibit significant homology to SCRs 2, 15 and 19 of human factor H, and SCRs 11, 12 and 13 have homology to SCRs 1, 2 and 3 of human C4b-binding protein. For the first time a complete cDNA representing a putative complement-regulatory protein which is structurally related to mammalian complement proteins has been isolated from a bony fish.