Characterisation and gene sequencing of mussel foot proteins from indigenous marine mussel, Perna perna (Linnaeus, 1758): Nutrient immobilization and struvite synthesis.

Abstract

Researchers are more attracted to mussels because of their magical adhesive strength on the wet substratum. This peculiar character of mussels is achieved by moisture resistant, biocompatible and biodegradable polyphenolic proteins called mussel adhesive proteins or mussel foot proteins (MAP or MFP). In mussels mainly six different adhesive proteins (mfp-1 to mfp-6) are reported and they contain unusual amino acid residues called 3,4-dihydroxyphenylalanine (DOPA), which are mainly involved in adhesion. The understanding of mussel adhesion exposes many opportunities in the area of science and medicine. Even though, mussels are considered as worst participants of biofouling, which make serious economic losses. They accumulated on ship hulls, underwater pipes and industrial equipment which lead to extensive damage. In India, two major species of mussel are present namely, Perna perna and Perna viridis. The present study focused on P. perna adhesive machinery, adhesive protein characterisation and also the synthesis of slow-released fertilizer by using P. perna shell and cow urine. Strong adhesion of the mussel is achieved by involvement of two parts called foot and byssus thread. The structural elucidation of mussel adhesive machinery helps to develop new antifouling strategies. The first chapter of this thesis deals with the structural elucidation of P. perna foot and byssus thread. The byssus thread contains three regions namely proximal thread, distal thread and plaque. The proximal and distal threads are morphologically distinct, in which gradient distribution of proteins was shown, leading to indestructible strength. Plaque is a leaf-like structure found attached to a substratum containing major polyphenolic adhesive proteins with the differential distribution. The polyphenolic protein staining of byssus thread using NBT revealed that high concentration of DOPA-rich proteins found in the adhesive region of plaque. Likewise, anatomically thread contains two parts cuticle and a core region. The cuticle is a hard protective part made up of mfp-1 and the core region contains collagens which provide elasticity and extensibility. The byssus thread's incredible thermal stability was exhibited by TGA analysis and secondary structure analysis using FTIR showed the byssus thread's strength. The elemental composition analysis indicates that mussel byssus thread can use as an excellent pollution monitoring specimen. The adhesive proteins have high commercial value because of their non-toxic property. Natural and recombinant adhesive proteins are now commercially available. The second chapter of the present thesis concentrated on designing a new extraction protocol for the natural extraction of adhesive proteins from P. perna. Here, a new protocol was developed and the presence of polyphenolic foot protein 1 (fp-1) in the sample was determined by acid urea electrophoresis and UV- Visible spectroscopy. The extracted proteins pH stability was determined by UV-Visible spectroscopy, adhesive strength study was measured using AFM and hydrophobicity was determined by contact angle measurement. The third chapter of the thesis focused on gene sequencing of adhesive protein and insilico characterisation of theobtained and insilico characterisation results highlighted that mussel foot protein 6 two variants act as protease inhibitors and had high antioxidant activity. The mussel shells are an excellent source of minerals and mineral deficiency is one of the hurdles in the farming system. The fourth chapter of the thesis dealing the synthesis of new slow-release fertilizer by using cow urine and mussel shell powder. The combination of cow urine and shell powder leads to the formation of struvite crystals which provide a more slow-releasing property to fertilizer and also rich plants essential macro and micronutrients. Mytilus edulis is the most studied mussel in the field of adhesion research followed by Mytilus californianus, Mytilus galloprovincialis and P. viridis. The studies from P. perna are very basic and not much progressive. Hence, the present research work is open to new informative parts of adhesion research. obtained protein. Two variants of P. perna foot protein 6 were obtained and insilico characterisation results highlighted that mussel foot protein 6 two variants act as protease inhibitors and had high antioxidant activity. The mussel shells are an excellent source of minerals and mineral deficiency is one of the hurdles in the farming system. The fourth chapter of the thesis dealing the synthesis of new slow-release fertilizer by using cow urine and mussel shell powder. The combination of cow urine and shell powder leads to the formation of struvite crystals which provide a more slow-releasing property to fertilizer and also rich plants essential macro and micronutrients. Mytilus edulis is the most studied mussel in the field of adhesion research followed by Mytilus californianus, Mytilus galloprovincialis and P. viridis. The studies from P. perna are very basic and not much progressive. Hence, the present research work is open to new informative parts of adhesion research.