title: Hydrolysis of amelogenin by matrix metalloprotease-20 accelerates mineralization in vitro.
abstract: In the following respects, tooth enamel is a unique tissue in the mammalian body: (a) it is the most mineralized and hardest tissue in it comprising up to 95 wt% of apatite; (b) its microstructure is dominated by parallel rods composed of bundles of 40-60 nm wide apatite crystals with aspect ratios reaching up to 1:10,000 and (c) not only does the protein matrix that gives rise to enamel guides the crystal growth, but it also conducts its own degradation and removal in parallel. Hence, when mimicking the process of amelogenesis in vitro, crystal growth has to be coupled to proteolytic digestion of the amelogenin assemblies that are known to play a pivotal role in conducting the proper crystal growth. Experimental settings based on controlled and programmable titration of amelogenin sols digested by means of MMP-20 with buffered calcium and phosphate solutions were employed to imitate the formation of elongated, plate-shaped crystals. Whilst amelogenin can act as a promoter of nucleation and crystal growth alone, in this study we show that proteolysis exerts an additional nucleation- and growth-promoting effect. Hydrolysis of full-length amelogenin by MMP-20 decreases the critical time needed for the protein and peptides to adhere and to cover the substrate. The formation and immobilization of a protein layer subsequently reduces the time for calcium phosphate crystallization. Coupling the proteolytic reaction to titration in the presence of 0.4 mg/ml rH174 has been shown to have the same effect on the crystal growth promotion as quadrupling the concentration of rH174 to 1.6 mg/ml. Controlling the rate and the extent of the proteolytic cleavage can thus be used to control the nucleation and growth rates in a protein-guided crystallization system.
authors.affiliation: Division of Biomaterials and Bioengineering, Department of Preventive and Restorative Dental Sciences, University of California, Parnassus Avenue 707, San Francisco, CA 94143, USA.
authors.forename: Vuk
authors.initials: V
authors.lastname: Uskoković
authors.forename: Feroz
authors.initials: F
authors.lastname: Khan
authors.forename: Haichuan
authors.initials: H
authors.lastname: Liu
authors.forename: Halina Ewa
authors.initials: HE
authors.lastname: Witkowska
authors.forename: Li
authors.initials: L
authors.lastname: Zhu
authors.forename: Wu
authors.initials: W
authors.lastname: Li
authors.forename: Stefan
authors.initials: S
authors.lastname: Habelitz
mesh_terms: D053523:Amelogenin / Q000737:chemistry / Q000378:metabolism*; D001031:Apatites / Q000737:chemistry / Q000378:metabolism*; D002130:Calcium Phosphates / Q000737:chemistry / Q000378:metabolism*; D003460:Crystallization; D003746:Dental Enamel Proteins / Q000737:chemistry / Q000378:metabolism*; D006801:Humans; D006868:Hydrolysis; D066298:In Vitro Techniques; D053519:Matrix Metalloproteinase 20 / Q000494:pharmacology*; D059748:Proteolysis; D013997:Time Factors
chemical_list: D053523:Amelogenin; D001031:Apatites; D002130:Calcium Phosphates; D003746:Dental Enamel Proteins; C020243:calcium phosphate; C507111:MMP20 protein, human; D053519:Matrix Metalloproteinase 20