How Much Do You Charge For N Acetyl L Cysteine > 자유게시판

• 목록
• 답변
• 글쓰기
• 게시판 리스트 옵션
  • 수정
  • 삭제

It has been discovered, N-Acetyl-L-Cysteine 98% manufacturers surprisingly, that the novel amino acid substitutions and/or amino acid deletions of the carboxyl terminus of the serine acetyltransferase result in a diminution in the cysteine sensitivity whereas at the identical time permitting satisfactory enzymic activity to be retained. It is feasible to cut back the cysteine sensitivity of the serine acetyltransferase in vivo by producing, by means of expression vectors, antisense RNAs which are complementary to a defined region of the 3' coding strand of the native or remodeled cysE gene. The gene for serine acetyltransferase has already been cloned and the amino acid sequence which is deduced from the DNA sequence is understood (Denk, D. and Bock, A. 1987, J. Gen. Microbiol. As well as, O-acetylserine sulfhydrylase B (cysM) is ready to make the most of thiosulfate as a sulfur source (Sirko, A. et al., 1987, J. Gen. Microbiol. Methods for introducing mutations at specific positions inside a DNA fragment are identified and are described, for example, in the next publications: Sarkar, G., Sommer, S. S., 1990, BioTechniques 8: 404-407 describe site-specific mutagenesis using PCR; Ausubel, F. M. et al., 1987, pp. Another method of producing feedback-resistant cysE alleles consists in combining totally different point mutations which lead to suggestions resistance, thereby giving rise to multiple mutants possessing new properties.

For this reason, the suggestions-resistant cysE alleles are ideally built-in into the genome as single copies using customary strategies. Strains which contain cysteine-sensitive proteins, for example prokaryotes or yeasts, are used as host strains. Since, in principle, cysteine metabolism proceeds by the use of the identical metabolic route, which is thought per se, in all microorganisms, and the techniques to be used for preparing the novel strains are well-known, for instance from commonplace textbooks, and relevant to all microorganisms, novel strains might be prepared from any microorganisms whatsoever. The invention also relates to the preparation of L-cysteine, or of merchandise that are derived from L-cysteine, via cultivating novel microorganisms. The invention additionally pertains to microorganisms which contain the feedback-resistant cysE alleles. The current invention furthermore relates to DNA sequences which encode novel serine acetyltransferases. The coding sequences that are present on the vector are advantageously linked to regulatory elements which are required for expressing the coding sequences to the desired extent. Sequences which encode selective markers and/or reporter genes are additionally ideally present on the expression vector in addition to the regulatory components.

An extra enhance in the cysteine yield may be achieved by additionally overexpressing the sulfate-reducing enzymes (encoded by the genes cysD, C, H, G, I and J) and the sulfhydrating enzymes (encoded by the genes cysK and cysM). The formation of L-cysteine itself is catalyzed by two O-acetylserine sulfhydrylase isoenzymes (EC 4.2.99.8), encoded by the genes cysK (O-acetylserine sulfhydrylase A) and cysM (O-acetylserine sulfhydrylase B), a response in which O-acetylserine features as a β-alanyl donor and H2 S as a β-alanyl acceptor (Kredich, N. M. and G. M. Tomkins 1966, J. Biol. The catalytic exercise of the completely different serine acetyltransferase enzymes is determined within the presence and absence of L-cysteine, and the inhibitor constant, Ki, is ascertained from this (Kredich and Tomkins, J. Biol. 1.1×10-6 M was determined within the presence of 0.1 mM acetyl-coenzyme A and 1 mM L-serine (Kredich, N. M. 1971 and Tomkins G. M. 1966, J. Biol.

The novel serine acetyltransferases ideally have an inhibitor constant, Ki, of from 0.005 to 2.Three mM within the presence of 1 mM L-serine and 0.1 mM acetyl-CoA, where serine acetyltransferases having not less than one mutation ideally possess an inhibitor fixed, Ki, of from 0.015 to 2.Three mM within the presence of 1 mM L-serine and 0.1 mM acetyl-CoA, whereas serine acetyltransferases having at the very least one carboxyterminal deletion preferably exhibit an inhibitor constant, Ki, of from 0.005 to 0.03 mM in the presence of 1 mM L-serine and 0.1 mM acetyl-CoA. Strategies for integrating genes into the chromosome utilizing vectors whose origins of replication have been eliminated are state of the art (Winans et al., 1985; J. Bacteriol. It is part of the state-of-the-art to block or modify gene exercise in a selected manner by means of so-called reverse genetics using antisense RNA (Inouye, 1988, Gene 72: 25-34). Antisense RNA is the transcription product of the DNA strand which is complementary to the strand encoding the protein. The starting DNA fragment, encompassing, for instance, the wild-type cysE gene, is recombined on a vector using known customary strategies for preparing recombinant DNA. The DNA of the wild-type cysE gene, or a cysE gene which has been inactivated by mutation, or a cysE gene which has been mutated and which already encodes a feedback-resistant serine acetyltransferase, is preferably used as the beginning material for the mutagenesis.