Blue Mussel Adhesive

The common blue mussel, Mytilus edulis, a marine delicacy savored by seafood lovers around the world, has a remarkable ability to cling to rocks, pilings and other underwater objects in turbulent tidal basins. Anyone who has harvested these shiny black mollusks knows how strong the attachments and the thin threads that tether them are and how difficult it is to remove those "beards" when preparing them for cooking. Scientists have long admired the mussel's perfect waterproof adhesive, but were unable to mimic its incredibly strong and durable properties. Researchers at the Idaho National Laboratory (INL) using reverse-genetics molecular biology techniques have assembled complete gene blueprints of the key, unique adhesive proteins secreted by the mussel. This breakthrough provides a new route to large-scale production of these adhesive proteins, potentially providing a "green" alternative to often toxic and expensive commercial adhesives used in everyday products from plywood to automobiles.

The INL blue mussel proteins may be used to develop an adhesive product that is capable of bonding  underwater, is impervious to water and turbulent forces and is environmentally safe, opening new applications in military, marine, and construction industries. Because such an adhesive could be biocompatible and moisture-proof, it could be used in a variety of biomedical and surgical applications as well.

Mussels like to position themselves in the fastest moving water and have adapted perfectly to their habitat by secreting quick-setting adhesive anchors and strong tethers (called byssal structures) from a foot organ in their shells. So far, scientists have identified nine protein components in the byssi. The proteins have specialized functions as primers, surface bonding promoters, adhesives, and catalysts (initiating an epoxy-like polymerization process).

Prior research shows that the most abundant protein in the adhesive, Mytilus edulis foot protein-1 (Mefp-1), bonds to glass, ceramic, wood, rock, concrete, plastic (even Teflon) and biological substrates including skin, muscle, bone and other tissues. Commercially available mussel adhesive is made using protein extraction techniques (10,000 mussels are needed to produce 1 gram of Mefp-1 protein!)  Protein extraction is neither environmentally friendly nor economically practical. No commercial product incorporates any of the other proteins that contribute to the exceptional underwater adhesion of the natural M. edulis mussel adhesive.

INL scientists realized that the complete gene sequences for Mefp-1, Mefp-2 (which plays a major role in forming the anchor plaques that bond to underwater objects) and the other proteins involved in byssi formation are necessary for mimicking the natural bio-adhesive. For the first time, using recombinant DNA techniques, INL scientists have mapped the nucleotide sequences from complementary DNA for Mefp-1, Mefp-2 and Mefp-3 as well as the full messenger RNA (ribonucleic acid) sequences for cloning. The isolation, purification and sequencing of these and other proteins by INL scientists are critical advances for replicating the natural blue mussel bio-adhesive and retaining its exceptional adhesion, strength and underwater properties. The development also opens a new, direct route to large-scale production of Mefp-1, Mefp-2 and other proteins through genetic engineering and recombinant DNA/cloning techniques. Using these techniques, low-value crops (such as tobacco or potato) can be converted into high-value crops containing blue mussel proteins. Simple processing could make commercial quantities of competitively priced, environmentally-friendly, blue mussel adhesive proteins available for the first time. It may provide a viable alternative for an industry that is actively seeking improved, "green," adhesives with properties similar to phenolics, epoxies, cyanoacrylates but not tied to expensive petrochemical feedstocks.

The INL blue mussel adhesives technology is the only approach that makes use of near full-length gene sequences with numerous amino acid repeats to express Mefp-1 and Mefp-2 proteins, the major components of the natural adhesives produced by blue mussels. This DNA sequencing/cloning technique provides the first direct route to large-scale production methods for producing commercial quantities of the adhesives. The only commercial product on the market is made by extracting Mefp-1 protein from mussels (10,000 mussels are needed to produce one gram of the protein). There is no commercial source of Mefp-2. Laboratory-prepared, extracted Mefp-1 protein is not as strong as the natural adhesive. The INL technology uses recombinant DNA techniques not protein extractionto mimic the natural adhesive in strength and other properties. Further, the INL technology makes it possible to use a range of hosts including plants, microorganisms, cell culture or animal pharma for large-scale production of both Mefp-1 and Mefp-2. Here is how the new INL technology improves on competitive products:

1.  NATURAL ADHESIVE

Mefp-1 and Mefp-2 are integral components of the blue mussels' adhesive protein complex that give the animal the ability to attach tenaciously to objects underwater. Mussel adhesive proteins are scleroproteins, which contribute mechanical strength to supporting structures in animals. Familiar scleroproteins include collagen, silk, elastin, fibroin, keratin, fibrin and resilin. Individual adhesive proteins formed and stockpiled in the foot organ of mussels are secreted to form attachment points (or plaques) and threads (called byssal structures). So far, scientists have identified at least nine protein components of the byssal structures, which have been designated Mefp-1, Mefp-2, Mefp-3, Mefp-4, etc. Natural adhesives derived from marine mussels have been considered for several decades as replacements for toxic, expensive and environmentally dangerous adhesives synthesized primarily from petroleum feedstocks. The challenge, until now, has been obtaining reliable, pure, large-scale supplies of the adhesive proteins. The INL work has changed that and will revolutionize the field of adhesives technology.

2. USES RECOMBINANT DNA TECHNIQUES NOT PROTEIN EXTRACTION  

Initial strategies for identifying the adhesive proteins of blue mussels involved extracting and isolating proteins directly from the byssi of thousands of animals – to gain a gram of adhesive required processing 10,000 animals. This approach is used to produce small quantities of very expensive Mefp-1 product sold today (BD Biosciences Clontech, under the trademark CELL-TAK, for attaching animal cells to plastic vessels in cell culture applications). This is neither environmentally friendly nor economically practical. When the original mussel adhesive protein was identified, recombinant protein techniques using synthetic gene constructs were tried but the resulting protein was not as strong as the natural adhesive. INL scientists realized that the complete gene sequence for Mefp-1, Mefp-2 and the other proteins involved in byssus formation is necessary for mimicking the natural bioadhesive. For the first time, using recombinant DNA techniques, INL scientists have uncovered the nucleotide sequence from a complementary DNA for Mefp-1 as well as the full messenger RNA (ribonucleic acid) sequence for cloning. Mefp-1, they have found, consists of 1,700 base pairs and 565 amino acids. The isolation, purification and sequencing of the Mefp-1, Mefp-2 and other proteins by INL scientists is a critical advance in replicating the natural bioadhesive and retaining its native adhesion, strength and underwater setup properties. It opens a new, direct route for large-scale production of Mefp-1, Mefp-2 and other proteins using genetic engineering and recombinant DNA/cloning techniques.

3. POTENTIAL TO PRODUCE LARGE QUANTITIES AT LOW COST

There are currently no large-scale commercial sources for Mefp-1 due to the high cost of extraction methods and inconsistencies in the quality of protein from earlier recombinant protein techniques using synthetic gene constructs. The reverse-genetics approaches used by INL researchers to obtain complete gene sequences, enzymatic screening of a complementary DNA library from the foot organ of M. edulis, and the use of DNA probes, provide the exact blueprint of the adhesive proteins expressed and transcribed by the mussel. With the complete gene sequences now available, the complementary DNA product is inserted (cloned) into a plasmid (a cellular element that exists and replicates autonomously in the cytoplasm). The expression plasmid then is coupled with alternate host systems (microorganisms, yeast, cells) to produce the adhesive proteins in quantities adequate for testing and analysis by the adhesives industry. This same approach also suggests a direct route to simple production methods to make recombinant adhesive proteins in low-value crops (such as tobacco or potato). These high-value crops would be harvested and processed to obtain commercial quantities of naturally expressed blue mussel adhesive proteins. Quantities of blue mussel adhesive available in pound rather than gram quantities at low prices will stimulate research and development in the adhesives industry and user community for a broad range of applications. As demand grows, the INL recombinant DNA/cloning technology will allow the conversion of low-value crops into high-value designer crops that produce natural, high-performance adhesive proteins at competitive prices.

The development of a biomimetic adhesive product – an adhesive that employs man-made materials to mimic the efficient attachment mechanisms and underwater adhesion of the natural mussel – will revolutionize the field of adhesive technology. Such an adhesive is a valuable asset to the military (for easy attachment of underwater surveillance, measurement and ordnance devices) and to a wide range of industries and biomedical applications. All can benefit from an environmentally safe, biocompatible, strong, inexpensive alternative to the conventional adhesives available today. There are no conventional glues that can be applied in an aqueous environment and are impervious to water and turbulent forces. 

Potential Applications include:

1. UNDERWATER ADHESIVES

Military: attachment of sensing devices to underwater objects or to animal agents (such a s dolphins); repairs to underwater structures and vessels; emergency repairs to SCUBA and other diving apparatus

Marine: construction of and repairs to vessels

Communications: splicing adhesive for undersea cables; on-site underwater cable and repeater repairs

Water treatment facilities: fabrication and repairs in wet environments in treatment plants

Home Water Systems: easier fabrication and repair of home system for contractors and do-it-yourselfers

 2. CONSTRUCTION AND SPECIALTY ADHESIVES

The mechanical properties of blue mussel adhesives compare favorably with phenolics, epoxies and cyanoacrylates, mainstays of the construction and specialty adhesives industries, but without their toxic, environmentally damaging side effects. Phenolic resins that glue together the multiple wood sheets in plywood represent a $2.3 billion market by 2006; cyanoacrylates – an estimated $3.3 billion market by 2007 – are growing fastest in on-site construction and medical applications. As adhesives manufacturers look for ways to produce "greener" products, lower costs and move away from feedstocks tied to rocketing crude oil prices, Blue mussel adhesives have the potential to replace these competitors in the following applications:  

Forest products industry: stronger bonding agent for manufacturing plywood, oriented-strand materials and other building materials that deteriorate when subjected to water and moisture.

Construction: new adhesives for plumbing, paneling, trim, glazing, cement, electrical and architectural treatments

Home repair: strong, quick-set adhesives for shower, bathtub and plumbing repair where moisture is a problem

General repair: new "super glue" that is environmentally safe (vs cyanoacrylates now used)

Electronics: adhere ferromagnetic materials to hard disk metal substrates

3. BIOMEDICAL ADHESIVES

Surgical: incision closure -- glue rather than stitch or staple (expected to be a $1.5 billion market by 2011)

Trauma: emergency wound closure for battlefield injuries, EMT and other first responders, home medicine cabinets

Cardiology: venous graft procedures; affixing and repairing implants such as pacemakers and defibrillators

Dentistry: better, safer adhesive for affixing dentures, implants and repairing broken teeth

Ophthalmic: corneal implant adhesive

Orthopedics: implant cement

 4. ADHESIVES FOR JOINING HARD-TO-BOND MATERIALS

Plastics such as acetals, polyethylenes, fluoropolymers, polypropylene and thermoplastic vulcanizers (TPVs) are used in virtually all industries because of their low cost and versatile design capabilities. Many assembly methods exist for joining similar and/or dissimilar plastics – from mechanical to chemical methods. Adhesive assembly, however, provides unique benefits such as the ability to bond and seal, even distribution of stress across a joint, rapid fixture and cure times, gap filling abilities and easy automation. Adhesive bonding of plastics is used in automotive components, industrial machines, plumbing, electronics, packaging, appliances and various other consumer items. The medical device industry has made hard-to-bond plastics, such as Teflon fluoropolymers, the materials of choice for a variety of devices.  But only a few industrial adhesives offer consistently high bond strengths on hard-to-bond plastics.

Research on the blue mussel adhesive proteins is helping to advance the field of biomimetics, demonstrating new recombinant techniques for mimicking natural substances. The technology opens potential new and exciting routes to everyday products that will be less energy intensive and more environmentally friendly.

New knowledge and a growing understanding of how blue mussel adhesives bind so tenaciously underwater to various substrates may also suggest ways to decrease or eliminate bond formation, and develop anti-fouling solutions for naval and commercial maritime vessels.