Isolation and initial characterisation of complement components C3 and C4 of the nurse shark and the channel catfish

Area of Science:

• Immunology
• Biochemistry
• Evolutionary Biology

Background:

• The complement system is crucial for innate and adaptive immunity.
• Complement components C3 and C4 are central to complement activation pathways.
• Understanding complement evolution across vertebrates provides insights into immune system development.

Purpose of the Study:

• To isolate and structurally characterize complement components C3 and C4 from cartilaginous (nurse shark) and bony (channel catfish) fish.
• To compare the structure and N-terminal sequences of fish C3 and C4 with those of higher vertebrates.
• To provide the first structural evidence for complement C4 in both major fish lineages.

Main Methods:

• Serum protein isolation and purification.
• Protein structural analysis (chain composition, molecular mass).
• N-terminal amino acid sequencing.

Main Results:

• Complement C3 and C4 were successfully isolated from nurse shark and channel catfish serum.
• Fish C4 proteins exhibited a three-chain structure, while C3 proteins had a two-chain structure, mirroring higher vertebrates.
• Intra-chain thioester bonds were identified in the largest polypeptides of all four proteins.
• N-terminal sequencing revealed sequence similarities to mammalian C3 and C4, with an exception for catfish C3 alpha-chain.
• Nurse shark C2n was confirmed as the analogue of mammalian C4.

Conclusions:

• This study provides the first structural evidence for complement C4 in both cartilaginous and bony fish.
• The conserved structural features of C3 and C4 suggest an ancient evolutionary origin for these key complement proteins.
• Findings contribute to understanding the evolutionary trajectory of the complement system across vertebrate classes.