SYNTHETIC SPIDER DRAGLINE SILK PROTEINS AND THEIR PRODUCTION IN ESCHERICHIA-COLI

Synthetic genes were designed to encode analogs of the two proteins of Nephila clavipes dragline silk, spidroins 1 and 2. The genes were con structed of tandem repeats of relatively long (more than 300 bp) DNA s equences assembled from synthetic oligonucleotides, and encoded protei ns of high molecular mass (65-163 kDa). Both analogs were produced eff iciently in Escherichia coli. The yield and homogeneity of the product s of longer genes were limited by premature termination of synthesis, probably as a result of processivity errors in protein synthesis. Aver age termination rates were determined to be 1 in 1100 codons to 1 in 3 00 codons, depending on the length and synonymous codon choices of the gene. Both analog proteins could be induced to form stable aqueous so lutions without denaturants. Circular dichroism spectra of the purifie d proteins in dilute solution resembled spectra of redissolved natural dragline silk in reflecting a largely disordered structure in water a nd more ordered structures in mixed solvents with methanol and trifluo roethanol.