Sequence of a partialcDNA clone from a major protein of Nephila clavipes dragline silk

Spider silk is an unusual biological material which has a tensile strength greater than steel and elasticity greater than wool. These specific properties are a consequence of the amino acid sequence and protein structure. Silks from different glands and different species do have different characteristics. As a first step toward understanding the relationship between the protein structure and fiber properties, and also to engineer silks with specific qualities, I studied one of the silk proteins.; The research has focused on the dragline silk protein from Nephila clavipes. The silk fibers were collected from the spigot of Nephila clavipes and the fiber proteins were fragmented. The fragments were purified by reverse phase HPLC on a C-18 column and then sequenced. DNA probes were synthesized based on a peptide, GAGQG, one of the peptide fragments purified from HPLC. Using pBluescript plasmid as vector, a cDNA library was generated from major ampullate silk gland mRNA. The library was screened with these probes. A cDNA clone with a 2.4 kbp insert was sequenced. This DNA encodes the carboxy terminal portion of a silk protein. A repetitive sequence of polyalanine alternating with a glycine rich region was found in this protein. The current model is that the elasticity of the protein stems from the polyalanine region which forms a helix under a stretching whereas the glycine rich region forms the b-sheet crystalline domain responsible for the tensile strength. There is a high bias of codon usage in this protein gene. A and T are chosen in the third position of the codons for most of amino acids. The GC content is 62% even with the A and T preference because of the high percentage of glycine and alanine. It is possible more than one protein may be involved in the dragline silk fiber formation and the molecular weight of the major protein may be between 150,000 and 300,000.