It has been found, N-Acetyl-L-Cysteine B2B suppliers surprisingly, that the novel amino acid substitutions and/or amino acid deletions of the carboxyl terminus of the serine acetyltransferase lead to a diminution in the cysteine sensitivity while at the identical time allowing adequate enzymic exercise to be retained. It is possible to cut back the cysteine sensitivity of the serine acetyltransferase in vivo by producing, by the use of expression vectors, antisense RNAs that are complementary to an outlined area of the 3' coding strand of the native or transformed 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 utilize thiosulfate as a sulfur source (Sirko, A. et al., 1987, J. Gen. Microbiol. Methods for introducing mutations at specific positions within a DNA fragment are known and are described, for example, in the next publications: Sarkar, G., Sommer, S. S., 1990, BioTechniques 8: 404-407 describe site-particular mutagenesis using PCR; Ausubel, F. M. et al., 1987, pp. Another method of producing feedback-resistant cysE alleles consists in combining completely different level mutations which lead to feedback resistance, thereby giving rise to a number of mutants possessing new properties.

For this reason, the suggestions-resistant cysE alleles are preferably integrated into the genome as single copies using customary strategies. Strains which include cysteine-delicate proteins, for example prokaryotes or yeasts, are used as host strains. Since, in precept, cysteine metabolism proceeds by means of the same metabolic route, which is thought per se, in all microorganisms, and the methods for use for making ready the novel strains are well-known, for example from normal textbooks, and applicable to all microorganisms, novel strains could be ready from any microorganisms in anyway. The invention additionally relates to the preparation of L-cysteine, or of merchandise which are derived from L-cysteine, via cultivating novel microorganisms. The invention additionally relates to microorganisms which comprise the suggestions-resistant cysE alleles. The present invention moreover relates to DNA sequences which encode novel serine acetyltransferases. The coding sequences that are present on the vector are advantageously linked to regulatory components which are required for expressing the coding sequences to the specified extent. Sequences which encode selective markers and/or reporter genes are additionally ideally current on the expression vector along with the regulatory elements.

A further improve in the cysteine yield could be achieved by moreover overexpressing the sulfate-lowering 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 reaction 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 activity of the completely different serine acetyltransferase enzymes is decided 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 decided in 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 in the presence of 1 mM L-serine and 0.1 mM acetyl-CoA, the place serine acetyltransferases having not less than one mutation preferably possess an inhibitor constant, Ki, of from 0.015 to 2.Three mM within the presence of 1 mM L-serine and 0.1 mM acetyl-CoA, while serine acetyltransferases having not less than one carboxyterminal deletion preferably exhibit an inhibitor fixed, 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 cutting-edge (Winans et al., 1985; J. Bacteriol. It's a part of the state-of-the-art to dam or modify gene exercise in a specific manner by the use 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 example, the wild-type cysE gene, is recombined on a vector using recognized customary techniques for preparing recombinant DNA. The DNA of the wild-sort 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 suggestions-resistant serine acetyltransferase, is ideally used as the starting material for the mutagenesis.