ENGINEERING TRIMERIC FIBROUS PROTEINS BASED ON BACTERIOPHAGE-T4 ADHESINS

The adsorption specificity of bacteriophage T4 is determined by genes 12 and 37, encoding the short tail-fibers (STF) and the distal part of the long tail-fibers (LTF), respectively Both are trimeric proteins w ith rod domains made up of similar tandem quasi-repeats, similar to 40 amino acids long. Their assembly requires the viral chaperones gp57A and gp38, Here we report that fusing fragments of gp12 and gp37 to ano ther trimeric T4 fibrous protein, fibritin, facilitates correct assemb ly, thereby by-passing the chaperone requirement. Fibritin is an alpha -helical coiled coil protein whose C-terminal part (fibritin E, compri sing the last 120 residues) has recently been solved to atomic resolut ion, Gp12 fragments of 109 and 70 amino acids, corresponding to three and two quasi-repeats respectively, were fused to the C-terminus of fi britin E. A similar chimera was designed for the last 63 residues of g p37, which contain four copies of the pentapeptide Gly-X-His-X-His and assume a narrow rigid structure in the LTF distal tip, Expressed from plasmids, ail three chimeras form soluble trimers that are resistant to dissociation by SDS and digestion by trypsin, indicative of correct folding and oligomerization.