{"id":{"repo_id":"aachen","oai_identifier":"oai:publications.rwth-aachen.de:61584"},"canonical_url":"https://search.dev.ndltd.org/etd/aachen/oai:publications.rwth-aachen.de:61584","repository":{"repo_id":"aachen","name":"RWTH Aachen University","base_url":"https://publications.rwth-aachen.de/oai2d"},"display":{"title":"Untersuchungen zur L-Serin-Bildung mit Corynebacterium glutamicum","abstract":"L-serine is a very important precursor of many cellular metabolites with a central position in the metabolism of corynebacterium glutamicum. This is especially because of the reaction catalyzed by the serine hydroxymethyltransferase (SHMT) in which serine is degraded to glycine and a C1 unit. It was an aim of this work to analyze the formation of the pharmaceutical important L-serine with metabolic engineering. Another aim was to analyze the regulation of the glyA-gene, coding for the SHMT.1. The wildtype of c. glutamicum does not accumulate L-serine and the overexpression of the serine pathway genes serA(fbr)CB only leads to a small increase in serine production to 0,05 mM. If the L-serine dehydratase is deleted, up to 3 mM L-serine is accumulated. With an insertion of the tac-promoter into the chromosome it was possible, to decrease the specific-activity of the essential SHMT from 30 to 1.3 nmol/min*mg protein and a very high production of 100 mM L-serine was achieved. Another further increase to 140 mM was gained by deleting the gene coding for the pyruvate kinase.2. The strain with glyA under control of the tac-promoter was instable, which was due to a mutation A13T in the LacIq-repressor protein. Because of this instability the genes for the folate-biosynthesis pabABC were deleted, because folate is needed for the synthesis of the SHMT-cofactor tetrahydrofolate. Strains with deleted pabABC-genes need 1 mM folate or p-aminobenzoate for growth. Folate-deficiency, overexpression of serA(fbr)CB and deletion of the L-serine dehydratase leaded to a very strong increase in L-serine production to 600 mM in a 20 l fermenter.3. The -10- and the -35-region of the glyA-gene could be determined by primer-extension-analysis. The transcriptional start site is 64 bp before the glyA-translational start site. Transcriptional fusions of the glyA-promoter with the chloramphenicol-acetyltransferase gene showed, that the glyA-promoter is – with 2 U/mg protein – a very strong promoter. With affinity chromatography two regulatory proteins were isolated which bind specifically to the promoter of the glyA-gene: ArsR and RamB. The ArsR-protein binds specifically to the glyA-gene in presence of 200 µM nickel(II) and enhances the glyA-expression by 20-25% in the stationary phase.","abstract_html":"L-serine is a very important precursor of many cellular metabolites with a central position in the metabolism of corynebacterium glutamicum. This is especially because of the reaction catalyzed by the serine hydroxymethyltransferase (SHMT) in which serine is degraded to glycine and a C1 unit. It was an aim of this work to analyze the formation of the pharmaceutical important L-serine with metabolic engineering. Another aim was to analyze the regulation of the glyA-gene, coding for the SHMT.1. The wildtype of c. glutamicum does not accumulate L-serine and the overexpression of the serine pathway genes serA(fbr)CB only leads to a small increase in serine production to 0,05 mM. If the L-serine dehydratase is deleted, up to 3 mM L-serine is accumulated. With an insertion of the tac-promoter into the chromosome it was possible, to decrease the specific-activity of the essential SHMT from 30 to 1.3 nmol/min*mg protein and a very high production of 100 mM L-serine was achieved. Another further increase to 140 mM was gained by deleting the gene coding for the pyruvate kinase.2. The strain with glyA under control of the tac-promoter was instable, which was due to a mutation A13T in the LacIq-repressor protein. Because of this instability the genes for the folate-biosynthesis pabABC were deleted, because folate is needed for the synthesis of the SHMT-cofactor tetrahydrofolate. Strains with deleted pabABC-genes need 1 mM folate or p-aminobenzoate for growth. Folate-deficiency, overexpression of serA(fbr)CB and deletion of the L-serine dehydratase leaded to a very strong increase in L-serine production to 600 mM in a 20 l fermenter.3. The -10- and the -35-region of the glyA-gene could be determined by primer-extension-analysis. The transcriptional start site is 64 bp before the glyA-translational start site. Transcriptional fusions of the glyA-promoter with the chloramphenicol-acetyltransferase gene showed, that the glyA-promoter is – with 2 U/mg protein – a very strong promoter. With affinity chromatography two regulatory proteins were isolated which bind specifically to the promoter of the glyA-gene: ArsR and RamB. The ArsR-protein binds specifically to the glyA-gene in presence of 200 µM nickel(II) and enhances the glyA-expression by 20-25% in the stationary phase.","abstract_has_math":false,"creators":["Stolz, Michael"],"institution":"Publikationsserver der RWTH Aachen University","degree_name":null,"degree_level":null,"degree_discipline":null,"degree_department":null,"school":null,"contributors":["Kreuzaler, Fritz"],"advisors":[],"committee_chairs":[],"committee_members":[],"year":2006,"date_issued":"2006","date_published":"2006","updated_at":"2026-07-30T19:43:10Z","subjects":["info:eu-repo/classification/ddc/570","Biowissenschaften, Biologie","L-Serin","Aminosäuren","SHMT, Produktion","L-serine","amino acids","SHMT, production"],"languages":["ger"],"rights":["info:eu-repo/semantics/openAccess"],"rights_urls":[],"identifier_entries":[{"key":"dc:identifier","label":"Identifier","values":["https://publications.rwth-aachen.de/search?p=id:%22RWTH-CONV-123235%22"],"render_values":[{"text":"https://publications.rwth-aachen.de/search?p=id:%22RWTH-CONV-123235%22","href":"https://publications.rwth-aachen.de/search?p=id:%22RWTH-CONV-123235%22","code":true}]}]},"links":{"outbound_url":"https://publications.rwth-aachen.de/record/61584","outbound_label":"Repository record","outbound_source":"dc:identifier"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor","label":"Contributor","values":["Kreuzaler, Fritz"]},{"key":"dc:creator","label":"Author","values":["Stolz, Michael"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:coverage","label":"Dc Coverage","values":["DE"]},{"key":"dc:date","label":"Dc Date","values":["2006"]},{"key":"dc:publisher","label":"Institution","values":["Publikationsserver der RWTH Aachen University"]},{"key":"dc:relation","label":"Dc Relation","values":["info:eu-repo/semantics/altIdentifier/urn/urn:nbn:de:hbz:82-opus-17453"]},{"key":"dc:type","label":"Dc Type","values":["info:eu-repo/semantics/doctoralThesis","info:eu-repo/semantics/publishedVersion"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["info:eu-repo/classification/ddc/570","Biowissenschaften, Biologie","L-Serin","Aminosäuren","SHMT, Produktion","L-serine","amino acids","SHMT, production"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:language","label":"Dc Language","values":["ger"]},{"key":"dc:rights","label":"Dc Rights","values":["info:eu-repo/semantics/openAccess"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier","label":"Identifier","values":["https://publications.rwth-aachen.de/record/61584","https://publications.rwth-aachen.de/search?p=id:%22RWTH-CONV-123235%22"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description","label":"Description","values":["L-serine is a very important precursor of many cellular metabolites with a central position in the metabolism of corynebacterium glutamicum. This is especially because of the reaction catalyzed by the serine hydroxymethyltransferase (SHMT) in which serine is degraded to glycine and a C1 unit. It was an aim of this work to analyze the formation of the pharmaceutical important L-serine with metabolic engineering. Another aim was to analyze the regulation of the glyA-gene, coding for the SHMT.1. The wildtype of c. glutamicum does not accumulate L-serine and the overexpression of the serine pathway genes serA(fbr)CB only leads to a small increase in serine production to 0,05 mM. If the L-serine dehydratase is deleted, up to 3 mM L-serine is accumulated. With an insertion of the tac-promoter into the chromosome it was possible, to decrease the specific-activity of the essential SHMT from 30 to 1.3 nmol/min*mg protein and a very high production of 100 mM L-serine was achieved. Another further increase to 140 mM was gained by deleting the gene coding for the pyruvate kinase.2. The strain with glyA under control of the tac-promoter was instable, which was due to a mutation A13T in the LacIq-repressor protein. Because of this instability the genes for the folate-biosynthesis pabABC were deleted, because folate is needed for the synthesis of the SHMT-cofactor tetrahydrofolate. Strains with deleted pabABC-genes need 1 mM folate or p-aminobenzoate for growth. Folate-deficiency, overexpression of serA(fbr)CB and deletion of the L-serine dehydratase leaded to a very strong increase in L-serine production to 600 mM in a 20 l fermenter.3. The -10- and the -35-region of the glyA-gene could be determined by primer-extension-analysis. The transcriptional start site is 64 bp before the glyA-translational start site. Transcriptional fusions of the glyA-promoter with the chloramphenicol-acetyltransferase gene showed, that the glyA-promoter is – with 2 U/mg protein – a very strong promoter. With affinity chromatography two regulatory proteins were isolated which bind specifically to the promoter of the glyA-gene: ArsR and RamB. The ArsR-protein binds specifically to the glyA-gene in presence of 200 µM nickel(II) and enhances the glyA-expression by 20-25% in the stationary phase."]},{"key":"dc:source","label":"Dc Source","values":["Aachen : Publikationsserver der RWTH Aachen University 100 S. : graph. Darst. (2006). = Aachen, Techn. Hochsch., Diss., 2006"]},{"key":"dc:title","label":"Title","values":["Untersuchungen zur L-Serin-Bildung mit Corynebacterium glutamicum"]}]}],"canonical_facts":{"dc:contributor":["Kreuzaler, Fritz"],"dc:coverage":["DE"],"dc:creator":["Stolz, Michael"],"dc:date":["2006"],"dc:description":["L-serine is a very important precursor of many cellular metabolites with a central position in the metabolism of corynebacterium glutamicum. This is especially because of the reaction catalyzed by the serine hydroxymethyltransferase (SHMT) in which serine is degraded to glycine and a C1 unit. It was an aim of this work to analyze the formation of the pharmaceutical important L-serine with metabolic engineering. Another aim was to analyze the regulation of the glyA-gene, coding for the SHMT.1. The wildtype of c. glutamicum does not accumulate L-serine and the overexpression of the serine pathway genes serA(fbr)CB only leads to a small increase in serine production to 0,05 mM. If the L-serine dehydratase is deleted, up to 3 mM L-serine is accumulated. With an insertion of the tac-promoter into the chromosome it was possible, to decrease the specific-activity of the essential SHMT from 30 to 1.3 nmol/min*mg protein and a very high production of 100 mM L-serine was achieved. Another further increase to 140 mM was gained by deleting the gene coding for the pyruvate kinase.2. The strain with glyA under control of the tac-promoter was instable, which was due to a mutation A13T in the LacIq-repressor protein. Because of this instability the genes for the folate-biosynthesis pabABC were deleted, because folate is needed for the synthesis of the SHMT-cofactor tetrahydrofolate. Strains with deleted pabABC-genes need 1 mM folate or p-aminobenzoate for growth. Folate-deficiency, overexpression of serA(fbr)CB and deletion of the L-serine dehydratase leaded to a very strong increase in L-serine production to 600 mM in a 20 l fermenter.3. The -10- and the -35-region of the glyA-gene could be determined by primer-extension-analysis. The transcriptional start site is 64 bp before the glyA-translational start site. Transcriptional fusions of the glyA-promoter with the chloramphenicol-acetyltransferase gene showed, that the glyA-promoter is – with 2 U/mg protein – a very strong promoter. With affinity chromatography two regulatory proteins were isolated which bind specifically to the promoter of the glyA-gene: ArsR and RamB. The ArsR-protein binds specifically to the glyA-gene in presence of 200 µM nickel(II) and enhances the glyA-expression by 20-25% in the stationary phase."],"dc:identifier":["https://publications.rwth-aachen.de/record/61584","https://publications.rwth-aachen.de/search?p=id:%22RWTH-CONV-123235%22"],"dc:language":["ger"],"dc:publisher":["Publikationsserver der RWTH Aachen University"],"dc:relation":["info:eu-repo/semantics/altIdentifier/urn/urn:nbn:de:hbz:82-opus-17453"],"dc:rights":["info:eu-repo/semantics/openAccess"],"dc:source":["Aachen : Publikationsserver der RWTH Aachen University 100 S. : graph. Darst. (2006). = Aachen, Techn. Hochsch., Diss., 2006"],"dc:subject":["info:eu-repo/classification/ddc/570","Biowissenschaften, Biologie","L-Serin","Aminosäuren","SHMT, Produktion","L-serine","amino acids","SHMT, production"],"dc:title":["Untersuchungen zur L-Serin-Bildung mit Corynebacterium glutamicum"],"dc:type":["info:eu-repo/semantics/doctoralThesis","info:eu-repo/semantics/publishedVersion"]},"updated_at":"2026-07-30T19:43:10Z"}