EP0260919A1
Expired / Lapsed
EP
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L-phenylalanine ammonia lyase
Assignee: Mitsui Chemicals Inc
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Filing: 1987-09-15
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Granted: —
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Expires: 2007-09-15
Abstract
Abstract In order to provide technical information necessary for the production of L-phenylalanine ammonia-lyase (PAL) by utilizing genetic engineering techniques, the structural gene for PAL and the amino acid sequence of PAL have been elucidated in Rhodosporidium toruloides , and novel recombinant DNA plasmids (e.g., pSW101, pYtrp6 and pKY201) have been created by inserting a DNA strand coding for the PAL gene between the 3ʹ-terminus of the promoter region and the 5ʹ-terminus of the terminator region. Moreover, transformants having such a novel recombinant DNA plasmid have been created, and a process for the production of PAL by growing such a novel transformant so as to cause PAL to be produced and accumulated in the culture has been established. Furthermore, there has been established a novel technique for the production of L-phenylalanine by reacting an ammonia donor with cinnamic acid in the presence of the PAL prepared by the aforesaid novel process.
Inventors
- 👤 Nobuhiro Fukuhara
- 👤 Setsuo Yoshino
- 👤 Kaoru Yamamoto
- 👤 Tomoyuki Se
- 👤 Satori Sone
- 👤 Yoshiyuki Mitsui Toatsu Kosugaya Nakajima
- 👤 Maki Mitsui Toatsu Iijima Apaato 2-47 Suzuki
- 👤 Nobuyoshi Makiguchi
Bibliographic Details
Application Number
—
Jurisdiction
EP
Data Source
google_patents
Last Synchronized
2026-11-05 17:24
CPC Classifications
C; C12; C12N; C12N15/00; C12N15/09; C12N15/11; C12N15/52; C12P; C12P13/00; C12P13/04; C12P13/22; C12P13/222; C12N9/00; C12N9/88
Claims Analysis
Total Claims: 16
Claim 1
Independent Claim
8 words
L-Phenylalanine ammonia-lyase having the following amino acid sequence:
Claim 2
Dependent Claim
8 words
L-Phenylalanine ammonia-lyase having the following amino acid sequence:
Claim 3
Dependent Claim
25 words
The DNA sequence functioning as a structural gene for L-phenylalanine ammonia-lyase and coding for the following amino acids 1 to 716 in the order mentioned:
Claim 4
Dependent Claim
25 words
The DNA sequence functioning as a structural gene for L-phenylalanine ammonia-lyase and coding for the following amino acids 1 to 716 in the order mentioned:
Claim 5
Dependent Claim
40 words
A DNA sequence coding for the amino acids 1 to 716 described in claim 2 in the order mentioned and functioning as a structural gene for L-phenylalanine ammonia-lyase in substantially the same manner as the DNA sequence of claim 2.
Claim 6
Dependent Claim
20 words
The DNA. sequence of claim 2 or 3 which has terminator codons at the C-terminus , e.g. TAG or TAA.
Claim 7
Dependent Claim
108 words
A process for the preparation of DNA coding for a structural gene for L-phenylalanine ammonia-lyase derived from Rhodsporidium toruloides which comprises the steps of (a) growing the yeast Phodosporidium toruloides under conditions capable of inducing L-phenylalanine ammonia-lyase, (b) separating a fraction containing the mRNA for L-phenylalanine ammonia-lyase from the yeast cells, (c) generating single-stranded cDNA from the mRNA by use of reverse transcriptase, (d) converting the single-stranded cDNA into double-stranded cDNA, (e) inserting the double-stranded cDNA into a vector, (f) transforming hose cells with the vector, (g) producing a library of cDNAs, and (h) cloning the cDNA coding for the structural gene for L-phenylalanine ammonialyase from the library.
Claim 8
Dependent Claim
108 words
A process for the preparation of DNA coding for a structural gene for L-phenylalanine ammonia-lyase derived from Rhodsporidium toruloides which comprises the steps of (a) growing the yeast Phodosporidium toruloides under conditions capable of inducing L-phenylalanine ammonia-lyase, (b) separating a fraction containing the mRNA for L-phenylalanine ammonia-lyase from the yeast cells, (c) generating single-stranded cDNA from the mRNA by use of reverse transcriptase, (d) converting the single-stranded cDNA into double-stranded cDNA, (e) inserting the double-stranded cDNA into a vector, (f) transforming hose cells with the vector, (g) producing a library of cDNAs, and (h) cloning the cDNA coding for the structural gene for L-phenylalanine ammonialyase from the library.
Claim 9
Dependent Claim
46 words
A recombinant DNA plasmid comprising an expression vector having incorporated therein a DNA strand whose DNA sequence codes for a polypeptide having the amino acid sequence described in claim 1, the expression vector having a construction permitting the polypeptide to be expressed in a desired host.
Claim 10
Dependent Claim
39 words
The recombinant DNA plasmid of claim 6 wherein the DNA strand is incorporated in the expression vector so as to be located between the 3ʹ-terminus of the promoter for the DNA sequence and the 5ʹ-terminus of the terminator therefor.
Claim 11
Dependent Claim
33 words
The recombinant DNA plasmid of claim 6 which is (a) a novel re combinant DNA plasmid pSW101; or (b) a novel recombinant DNA plasmid pYtrp6; or (c) a novel recombinant DNA plasmid pKY201.
Claim 12
Dependent Claim
19 words
A recombinant microorganism productive of L-phenylalanine ammonia-lyase which is a transformant having the recombinant DNA plasmid of claim 6.
Claim 13
Dependent Claim
39 words
The recombinant microorganism of claim 9 which is (a) a novel transformant FERM P-8834 of Escherichia coli ; or (b) a novel transformant FERM P-8876 of Escherichia coli ; or (c) a novel transformant FERM P-8875 of bakers' yeast.
Claim 14
Dependent Claim
34 words
A process for the production of L-phenylalanine ammonia-lyase which comprises growing the recombinant microorganism of claim 9 or claim 10 so as to cause L-phenylalanine ammonia-lyase to be produced and accumulated in the culture.
Claim 15
Dependent Claim
15 words
L-phenylalanine ammonia-lyase which is produced by a recombinant microorganism of claim 9 or claim 10.
Claim 16
Dependent Claim
52 words
A process for the production of L-phenylalanine which comprises the steps of growing the recombinant microorganism of claim 9 or claim 10 so as to cause L-phenylalanine ammonia-lyase to be produced and accumulated in the culture, and reacting an ammonia donor with cinnamic acid in the presence of the produced L-phenylalanine ammonia-lyase.
References Cited (Backward Citations)
EP0137280A1
Cited
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