Mussel decapeptide

The polyphenolic adhesive protein of the mussel Mytilus edulis is an unusual protein composed mainly of repetitive decapeptide and hexapeptide sequences. In the mussel, the protein is first produced in a precursor form and is converted to an adhesive by post-translation-al modification. To develop an efficient renewable resource for production of the polyphenolic protein, we have used genetic engineering technology. cDNA sequences encoding portions of the polyphenolic protein were identified and expressed in the yeast Saccharomyces cerevisiae. 

The development of a technology for the biological and chemical synthesis of analogs to the adhesive protein of the mussel Mytilus edulis is described. This protein consists mainly of the repeating decapeptide sequence Ala-Lys-Pro-Ser-Tyr-Hyp-Hyp-Thr-Tyr-Lys. 

The sequence of oxidation steps which converts the mussel adhesive protein to underwater glue was investigated using model peptides. The three-dimensional structure of the protein was derived from the decapeptide sequence with the help of computer modeling studies. The oxidation chemistries were then applied to the three-dimensional protein structure to develop a mechanistic picture, at the molecular level, showing how the mussel attaches itself to solid surfaces. 

Structural proteins such as collagen, elastin, and spider silk contain repeating sequences of amino acids. In 1983, Waite discovered that the protein secreted by the phenol gland of the blue sea mussel Mytilus edulis consists of closely related decapeptide and hexapeptide sequences with a combined molecular weight of approximately 130 kilodaltons (kDa).i He further elucidated that this protein is transformed into glue through enzymatic oxidation to form the adhesive plaques that anchor the mussel... 

In this paper, we report our progress in synthesizing the mussel adhesive protein and developing a mechanistic model for its conversion to a water-compatible glue. Following the EXPERIMENTAL section the results are presented in three sections under RESULTS AND DISCUSSION (1) Production of a Synthetic Mussel Adhesive Protein in Bacterial Systems, (2) Chemical Synthesis and Polymerization of Decapeptides, and (3) Mechanism of Glue Formation. 

Our research has centered on the production of a mussel glue analog protein in bacteria. The primary objective was the practical implementation of a gene cassette approach to the biological preparation of adhesive polymers. A synthetic DNA sequence has been constructed which codes for tandem repeats of the decapeptide precursor. This sequence has been cloned into E, coli. Our results show that a polydecapeptide analog protein of 25 kDa was successfully expressed from the synthetic gene. 

We studied the enzymatic oxidation of tyrosine using the decapeptide with the consensus sequence as the substrate. Since catechol oxidase from sea mussels is not available, we instead used mushroom tyrosinase for our studies. The oxidations were carried out in a phosphate buffer at ambient temperatures. The reactions were followed using UV spectroscopy and products were separated by HPLC and identified by FAB/MS. The oxidations were also followed in an NMR tube using D2O as the solvent. When the decapeptide Glue-2 (see Table 1) with the consensus sequence was oxidized in the presence of ascorbic acid, mono-dopa was the predominant product. The enzyme selectively oxidized the tyrosine at position 9.20 Glue-7, in... 

The chemical reactions discussed above are not by themselves sufficient to develop a mechanistic scheme for underwater glue formation unless they are reviewed in the context of the three-dimensional structure of the mussel adhesive protein. The structure of this protein however is not available. We therefore decided to derive its three-dimensional structure from the consensus decapeptide sequence in an incremental fashion using computer modeling and chemical intuition. It is well-known that the primary amino acid sequence determines the structure of a protein. However, predicting three-dimensional structures from an amino acid sequence is a risky venture unless there are experimental guide posts that support or reject the predicted structures. 

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