| The underwater adhesive ability of marine organisms in wet environments is one of the goals of underwater biomimicry.Many marine organisms(such as mussels,barnacles and sandcastle worms)have strong underwater adhesion abilities.The design and development of artificial underwater adhesion materials by mimicking their adhesion mechanisms on wet surfaces is an important way to solve the problems of bonding underwater equipment and devices.The mussel foot proteins from the marine mussel are the most studied underwater adhesion materials,and the 3,4-dihydroxyphenylalanine(DOPA),which is abundant in mussel foot proteins,has played key roles in mussel adhesion strength.A variety of adhesion materials have been developed based on the adhesion mechanism of mussel foot proteins,which can achieve adhesion in a variety of wet environments.Creatures among microorganisms also have the ability to adhere underwater,Caulobacter crescentus secretes a sticky polysaccharide called holdfast which has higher adhesion than that of chemical adhesives.It has an adhesion strength of68 Mpa under microscopic conditions,which is the highest known natural adhesive substance.The holdfast polysaccharide with N-acetylglucosamine as the basic unit helps the C.crescentus to colonise different material surfaces in turbulent currents,but the mechanism of its adhesion to the surface remains to be investigated.The study of mussel foot proteins and C.crescentus holdfast will help to unravel the mechanisms of adhesion of marine organisms in wet environments and to design and develop more adaptable biomimetic adhesion materials.The aim of this dissertation was to provide a theoretical basis for the development of bionic adhesion materials by analyzing the structure and adhesion mechanism of mussel foot protein and Caulobacter crescentus holdfast.Based on the results,the DOPA modification of polyglucosamine-like polysaccharide(chitosan),which was a holdfastlike material,was used to try out the novel biomimetic adhesion materials by combining the adhesion properties of the above two marine organisms.In this dissertation,we designed and synthesized different amino acid and DOPA pair structures to modify onto atomic force microscope(AFM)probes.Their adhesion and cohesion properties were tested and analyzed at the cation-rich(mica)and cationdeficient(modified silicon nitride,Si3N4(OH))interfaces by using single-molecule force spectroscopy(SMFS).Then the heterogeneous distribution of peptides’ amino acid residues was investigated by means of short peptide sequences intercepted from the mussel foot protein Mfp-3.The holdfast polysaccharide was enriched and extracted by using magnetic microspheres culturing with Caulobacter crescentus,and the structural characteristics and adhesion properties of holdfast were characterized by using atomic force microscopy.Although holdfast,a natural polysaccharide with super-adhesive properties,which is secreted in small amounts and difficult to extract,substances similar to its composition can be found in nature.Chitin,which also has a basic unit of Nacetylglucosamine,is widely distributed in the marine.Chitosan as its deacetylation product is abundantly produced and has similar characteristics to holdfast in producing adhesion by deacetylation.Therefore,chitosan with holdfast-like composition was investigated in terms of solubility,viscosity,and adhesion to analyze the effects of its molecular weight and degree of deacetylation on adhesion properties.In the previous work,we expressed the key enzyme-deacetylase from Caulobacter crescentus that induced holdfast adhesive.In order to mimic the mechanism of holdfast production,we optimized the activity of recombinant deacetylase and tried to modify chitosan with low deacetylation degree by recombinant deacetylase to make chitosan obtaining holdfastlike adhesive properties.Finally,by combining the adhesion properties of two natural adhesive materials,mussel foot protein and C.crescentus holdfast,the carboxylated chitosan was grafted by DOPA to retain its free amino group for protonation.The grafting of DOPA onto chitosan formed an adhesion mechanism similar to that of DOPA-Lys to the interface.The cohesion and adhesion of materials are equally important for their performance.Natural spider silk has excellent mechanical properties such as high strength,high tenacity,and low density.The nanoprimary fiber structure of spider silk protein in its composition provides strong tensile strength.To improve the cohesion of materials,recombinant spider silk protein was utilized as a fiber backbone structure to provide cohesive strength,and assembled with polydopamine(PDA)to form protein copolymers.The recombinant spider silk protein-PDA copolymer was screened for concentration ratios,and the polydopamine was cured by Fe Cl3 and HCl.The recombinant spider silk protein with fibrous structure and the polydopamine with cured mesh structure were cross-linked with each other,which could further enhance the cohesive properties of the copolymers.It was proposed that the self-assembled skeleton structure of recombinant spider silk protein strengthened the cohesive ability of the material.By combining with the interfacial adhesion ability of bidentate hydrogen bonding of polydopamine,the cohesive copolymer with both cohesive and adhesion strengths was developed.The results showed that the protonated amine in the DOPA-Lys pair structure could assist DOPA to form adjustable bidentate hydrogen bonds with cation-rich interfaces(mica)and cation-deficient interfaces(Si3N4(OH)).The multi-adaptive adhesion characteristics of Phe-Lys and DOPA-Trp were found to adhere to both types of interfaces via cation-π interactions and hydrophobic forces.The heterogeneous amino acid distribution and DOPA content in the polypeptide of the Mfp-3 fragment of the mussel foot protein played an important role in adhesion stability.Genome resequencing and physiological characterization of C.crescentus were determined.Based on the mechanism of post-contact stimulate secretion,enrichment and extraction of the less secreted,sticky holdfast was achieved by using polystyrene magnetic microspheres with particle sizes of 20 μm and 40 μm.Mechanical tests of holdfast using atomic force microscopy under air and underwater conditions yielded a maximum adhesion of 12.4 n N and 19.6 Nn to the Si3N4 probe tip,respectively.Results showed a stronger adhesion of holdfast with gel properties in the underwater environment.Chitosan,as a rare cationic polysaccharide,had the same composition and modification as holdfast.Its molecular weight was positively correlated with viscosity,and the degree of deacetylation was not closely related to viscosity,but closely correlated with adhesion strength.Activation of carboxylated chitosan carboxyl groups using EDC/NHS and grafting with DOPA could retain the free amino group in chitosan for the formation of protonated amine to assist DOPA adhesion.This modification method was trying to improve the adhesion strength of chitosan through a mechanism similar to the DOPA-Lys interaction.Grafting of DOPA onto chitosan enhanced the adhesion performance of chitosan on the surfaces of three materials(plastic,glass and wood).Analysis of the adhesion mechanism based on the surface properties of different materials showed that the bidentate hydrogen bonds of DOPA played an important role in enhancing the adhesion performance.Combining the high cohesive property of spider silk protein and the high adhesion property of mussel foot protein,the copolymer with recombinant spider silk protein and polydopamine showed that the copolymer solution had a high adhesion strength(2.64 MPa)on the surface of glass materials.The copolymer was formulated with 8.5% recombinant spider silk protein,0.2 M polydopamine and 0.03 M Fe Cl3 curing agent.The copolymer materials were characterized by using Fourier transform attenuated total reflection infrared spectroscopy(ATR-FTIR),water contact angle testing,and atomic force microscopy,respectively.The restoration of bidentate hydrogen bonds in the copolymer by 0.55 M HCl curing was found to improve the adhesion to silicon nitride probes(88n N)in the atomic force microscopy-based adhesion performance test.In conclusion,the adhesion mechanism of DOPA was investigated by designing and synthesizing dipeptides based on mussel foot protein,the structure of C crescentus holdfast was characterized by atomic force microscopy,which provided a theoretical basis for the design and development of new bionanomorphic adhesion materials.The chitosan-DOPA based on two marine organisms also successfully enhanced the adhesion performance of chitosan on different material surfaces,which provides ideas for further development into applicable adhesion materials.The copolymer of recombinant spieder silk protein and polydopamine,which were assembled by using two properties of high strength of spider silk proteins and high adhesion of mussel foot proteins,showed an excellent adhesion performance and provided an example for the development of novel protein copolymer adhesion materials. |
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