Chemistry: molecular biology and microbiology – Micro-organism – tissue cell culture or enzyme using process... – Recombinant dna technique included in method of making a...
Patent
1998-01-13
1999-10-19
Carlson, Karen Cochrane
Chemistry: molecular biology and microbiology
Micro-organism, tissue cell culture or enzyme using process...
Recombinant dna technique included in method of making a...
536 231, 530350, C12P 2106, C07H 1700, C07K 1400
Patent
active
059687720
DESCRIPTION:
BRIEF SUMMARY
This application has been filed under 35 U.S.C. 371 from PCT/JP96/01236, filed May 9, 1996, which claims priority to Japanese Patent Application 7-110877, filed May 9, 1995.
TECHNICAL FIELD
The present invention relates to proteins of constituting the pearl (pearl proteins), especially, to nacrein proteins produced through the genetic engineering techniques, and to a process for producing and expressing the proteins.
BACKGROUND ART
Shell of the pearl layer in the pearl oyster is consisting of minor proteinic organic matrix and crystal of aragonite. This organic matrix had been designated as "Conchiolin" (Fremy, M. E., Recherches chimiques sur les os. Ann. Chem. Phys, 43, 96 (1855)) and have been studied by many persons since then in all its aspects.
See, for example, the publications of: calcium-binding conchiolin shell peptides from the mollusc, Haliotis rufescens, as a function of development", J. Comp. Physiol. B, 157, pp. 717-729 (1988); Malacologia 22, pp. 225-233 (1981); different types of ultrastructure", The Veliger 33, pp. 190-201 (1990); mollusk shells: a potential template for shell formation", Science, 190, pp. 987-989 (1975); the organic matrices of mollusc shells", J. exp. mar. Biol. Ecol., 30, pp. 45-51 (1977); soluble organic matrix of mollusc shells", Calicif. Tissue Int., 29, pp. 163-167 (1979); and proteins associated with calcite deposition in the bivalve, Mytilus calfornianus", Biochemistry, 22, pp. 4139-4145 (1983).
Up to date, although it was reported that one of the main components in the organic matrix of the pearl layer is asparatic acid rich acidic protein in the molecular weight of 50.about.70 kD [See, publications (3) and (7) listed above], its accurate molecular weight and the amino acid sequence thereof have not been reported at all.
Accordingly, as a popular technology for forming pearl wherein a part of shell of Pinctada fucata is coaggulated, cultivation methods depending on environmental (cultural) conditions have been employed over one century or more to cultivate Pinctada fucata for one to two year(s) in a hanging sack in the sea.
DISCLOSURE OF INVENTION
The present invention is established to solve the problems noted above. That is to say, in addition to the genetical study/analysis of the pearl proteins, it is aimed to artificially produce the pearl proteins based on the unique genetic information of such proteins.
At the first step, the inventor extracted the substantial proteins from the natural pearl, purified them and isolated an acidic protein (nacrein) in the molecular weight of approximately 60,000 which is one of the major elements of the proteins and have Ca.sup.2+ (calcium ion) binding activity. Then, the N-terminal amino acids sequence of this protein was determined, then DNA probes corresponding to the sequence was synthesized.
On the other hand, cDNA library was constructed by extracting mRNA from mantle tissues of Pinctada fucata involved with production of the natural pearl layer. cDNA of nacrein was cloned from this library by a hybridization technique utilizing the synthesized DNA probes noted above.
Then, the nacrein was produced according to the known technique by linking cDNA so obtained to a gene expression vector and introducing the vector into host cell like E. coli. Watabe and Wilbur had already reported that the crystal of aragonite would be specifically produced on the organic matrix of the pearl layer (Watabe N. and Wilbur K. N., "Influence of the organic matrix on crystal type in Molluscs", Nature 188, 334 (1960)). Further, in view of the facts that the nacrein is one of the essential elements of the organic matrix and is the protein having Ca.sup.2+ -binding activity, in vitro production of the pearl layer will be realized by crystallizing the calcium carbonite to aragonite on the protein.
BRIEF DESCRIPTION OF DRAWINGS
FIG. 1 is a scheme showing an outline for extraction, concentration and purification of the nacrein proteins.
FIG. 2 is a graph showing an elution curve of the nacrein proteins obtained by DEAE-Sephacel Column Ch
LandOfFree
Pearl protein (nacrein) and process for producing the same does not yet have a rating. At this time, there are no reviews or comments for this patent.
If you have personal experience with Pearl protein (nacrein) and process for producing the same, we encourage you to share that experience with our LandOfFree.com community. Your opinion is very important and Pearl protein (nacrein) and process for producing the same will most certainly appreciate the feedback.
Profile ID: LFUS-PAI-O-2054635