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Biosynthesis of radiolabeled cellodextrins by the Clostridium thermocellum cellobiose and cellodextrin phosphorylases for measurement of intracellular sugars
The Clostridium thermocellum cellobiose and cellodextrin phosphorylases (glucosyl transferases) in the cell extract were used to synthesize radiolabeled cellodextrins with a degree of polymerization (DP=2-6) from nonradioactive glucose-1-phosphate and radioactive glucose. Chain lengths of synthesize...
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| Published in: | Applied microbiology and biotechnology 2006-03, Vol.70 (1), p.123-129 |
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| container_title | Applied microbiology and biotechnology |
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| creator | Zhang, Y. -H. P Lynd, L. R |
| description | The Clostridium thermocellum cellobiose and cellodextrin phosphorylases (glucosyl transferases) in the cell extract were used to synthesize radiolabeled cellodextrins with a degree of polymerization (DP=2-6) from nonradioactive glucose-1-phosphate and radioactive glucose. Chain lengths of synthesized cellodextrin were controlled by the absence or presence of dithiothreitol and by reaction conditions. All cellodextrins have the sole radioactive glucose unit located at the reducing ends. Mixed cellodextrins (G2-G6) were separated efficiently by size-exclusion chromatography or less efficiently by thin-layer chromatography. A new rapid sampling device was developed using disposable syringes containing an ultracold methanol-quenching buffer. It was simple, less costly, and especially convenient for anaerobic fermentation. After an impulse feed of radiolabeled cellobiose, the intracellular sugar levels were measured after a series of operations--sampling, extracting, concentrating, separating, and reading. Results showed that the largest amount of radioactivity was cellobiose with lesser amounts of glucose, cellotriose, and cellotetraose, and an average DP of intracellular cellodextrins was ca. 2. |
| doi_str_mv | 10.1007/s00253-005-0278-1 |
| format | article |
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P ; Lynd, L. R</creator><creatorcontrib>Zhang, Y. -H. P ; Lynd, L. R</creatorcontrib><description>The Clostridium thermocellum cellobiose and cellodextrin phosphorylases (glucosyl transferases) in the cell extract were used to synthesize radiolabeled cellodextrins with a degree of polymerization (DP=2-6) from nonradioactive glucose-1-phosphate and radioactive glucose. Chain lengths of synthesized cellodextrin were controlled by the absence or presence of dithiothreitol and by reaction conditions. All cellodextrins have the sole radioactive glucose unit located at the reducing ends. Mixed cellodextrins (G2-G6) were separated efficiently by size-exclusion chromatography or less efficiently by thin-layer chromatography. A new rapid sampling device was developed using disposable syringes containing an ultracold methanol-quenching buffer. It was simple, less costly, and especially convenient for anaerobic fermentation. After an impulse feed of radiolabeled cellobiose, the intracellular sugar levels were measured after a series of operations--sampling, extracting, concentrating, separating, and reading. Results showed that the largest amount of radioactivity was cellobiose with lesser amounts of glucose, cellotriose, and cellotetraose, and an average DP of intracellular cellodextrins was ca. 2.</description><identifier>ISSN: 0175-7598</identifier><identifier>EISSN: 1432-0614</identifier><identifier>DOI: 10.1007/s00253-005-0278-1</identifier><identifier>PMID: 16402169</identifier><identifier>CODEN: AMBIDG</identifier><language>eng</language><publisher>Berlin: Berlin/Heidelberg : Springer-Verlag</publisher><subject>Anaerobiosis ; Bacteriology ; Biological and medical sciences ; Biosynthesis ; Biotechnology ; Carbohydrate Conformation ; carbohydrate metabolism ; Carbon Radioisotopes ; cellobiose ; Cellulose - analogs & derivatives ; Cellulose - biosynthesis ; Cellulose - chemistry ; Cellulose - metabolism ; Chromatography ; Clostridium thermocellum ; Clostridium thermocellum - enzymology ; dextrins ; Dextrins - biosynthesis ; Dextrins - chemistry ; Dextrins - metabolism ; enzyme activity ; Fermentation ; Fundamental and applied biological sciences. Psychology ; Glucose ; Glucosyltransferases - metabolism ; hexosyltransferases ; Microbiology ; Radioactivity ; radiolabeling ; Sugar ; sugars ; Syringes ; Thin layer chromatography</subject><ispartof>Applied microbiology and biotechnology, 2006-03, Vol.70 (1), p.123-129</ispartof><rights>2006 INIST-CNRS</rights><rights>Springer-Verlag 2006</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c508t-6cef8f5bb2a838e4986531e017335d7c0fba9c37fe1d5fad69bc0794ab75217a3</citedby><cites>FETCH-LOGICAL-c508t-6cef8f5bb2a838e4986531e017335d7c0fba9c37fe1d5fad69bc0794ab75217a3</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.proquest.com/docview/625681150/fulltextPDF?pq-origsite=primo$$EPDF$$P50$$Gproquest$$H</linktopdf><linktohtml>$$Uhttps://www.proquest.com/docview/625681150?pq-origsite=primo$$EHTML$$P50$$Gproquest$$H</linktohtml><link.rule.ids>314,780,784,11688,27924,27925,36060,36061,44363,74895</link.rule.ids><backlink>$$Uhttp://pascal-francis.inist.fr/vibad/index.php?action=getRecordDetail&idt=17534550$$DView record in Pascal Francis$$Hfree_for_read</backlink><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/16402169$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Zhang, Y. -H. P</creatorcontrib><creatorcontrib>Lynd, L. R</creatorcontrib><title>Biosynthesis of radiolabeled cellodextrins by the Clostridium thermocellum cellobiose and cellodextrin phosphorylases for measurement of intracellular sugars</title><title>Applied microbiology and biotechnology</title><addtitle>Appl Microbiol Biotechnol</addtitle><description>The Clostridium thermocellum cellobiose and cellodextrin phosphorylases (glucosyl transferases) in the cell extract were used to synthesize radiolabeled cellodextrins with a degree of polymerization (DP=2-6) from nonradioactive glucose-1-phosphate and radioactive glucose. Chain lengths of synthesized cellodextrin were controlled by the absence or presence of dithiothreitol and by reaction conditions. All cellodextrins have the sole radioactive glucose unit located at the reducing ends. Mixed cellodextrins (G2-G6) were separated efficiently by size-exclusion chromatography or less efficiently by thin-layer chromatography. A new rapid sampling device was developed using disposable syringes containing an ultracold methanol-quenching buffer. It was simple, less costly, and especially convenient for anaerobic fermentation. After an impulse feed of radiolabeled cellobiose, the intracellular sugar levels were measured after a series of operations--sampling, extracting, concentrating, separating, and reading. Results showed that the largest amount of radioactivity was cellobiose with lesser amounts of glucose, cellotriose, and cellotetraose, and an average DP of intracellular cellodextrins was ca. 2.</description><subject>Anaerobiosis</subject><subject>Bacteriology</subject><subject>Biological and medical sciences</subject><subject>Biosynthesis</subject><subject>Biotechnology</subject><subject>Carbohydrate Conformation</subject><subject>carbohydrate metabolism</subject><subject>Carbon Radioisotopes</subject><subject>cellobiose</subject><subject>Cellulose - analogs & derivatives</subject><subject>Cellulose - biosynthesis</subject><subject>Cellulose - chemistry</subject><subject>Cellulose - metabolism</subject><subject>Chromatography</subject><subject>Clostridium thermocellum</subject><subject>Clostridium thermocellum - enzymology</subject><subject>dextrins</subject><subject>Dextrins - biosynthesis</subject><subject>Dextrins - chemistry</subject><subject>Dextrins - metabolism</subject><subject>enzyme activity</subject><subject>Fermentation</subject><subject>Fundamental and applied biological sciences. Psychology</subject><subject>Glucose</subject><subject>Glucosyltransferases - metabolism</subject><subject>hexosyltransferases</subject><subject>Microbiology</subject><subject>Radioactivity</subject><subject>radiolabeling</subject><subject>Sugar</subject><subject>sugars</subject><subject>Syringes</subject><subject>Thin layer chromatography</subject><issn>0175-7598</issn><issn>1432-0614</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2006</creationdate><recordtype>article</recordtype><sourceid>M0C</sourceid><recordid>eNqNkk1v1DAQhiMEokvhB3ABC6ncAjN2_JFjWZUPqRIH6NlyErtNlcSLJ5HYH8N_xdldqYILHCxrrOd9x6N5i-IlwjsE0O8JgEtRAsgSuDYlPio2WAlegsLqcbEB1LLUsjZnxTOiewDkRqmnxRmqCjiqelP8-tBH2k_znaeeWAwsua6Pg2v84DvW-mGInf85p34i1uxZ5th2iJQfun4Z1zqNccVycaCb7OeZm_4Us91dpHzSfnDkiYWY2OgdLcmPfprXxv00J3dwGlxitNy6RM-LJ8EN5F-c7vPi5uPV9-3n8vrrpy_by-uylWDmUrU-mCCbhjsjjK9qo6RAn8cXQna6hdC4uhU6eOxkcJ2qmxZ0XblGS47aifPi7dF3l-KPxdNsx57Wv7jJx4Ws0qqq0VT_BLGWXCKq_wB5nXdYZ_DNX-B9XNKUp7WKS2UQJWQIj1CbIlHywe5SP7q0twh2jYI9RsHmKNg1Chaz5tXJeGlG3z0oTrvPwMUJcNS6ISQ3tT09cFqKSh6avz5ywUXrblNmbr5xQAFgVCWNFL8BTLfJaw</recordid><startdate>20060301</startdate><enddate>20060301</enddate><creator>Zhang, Y. -H. 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P ; Lynd, L. R</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c508t-6cef8f5bb2a838e4986531e017335d7c0fba9c37fe1d5fad69bc0794ab75217a3</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2006</creationdate><topic>Anaerobiosis</topic><topic>Bacteriology</topic><topic>Biological and medical sciences</topic><topic>Biosynthesis</topic><topic>Biotechnology</topic><topic>Carbohydrate Conformation</topic><topic>carbohydrate metabolism</topic><topic>Carbon Radioisotopes</topic><topic>cellobiose</topic><topic>Cellulose - analogs & derivatives</topic><topic>Cellulose - biosynthesis</topic><topic>Cellulose - chemistry</topic><topic>Cellulose - metabolism</topic><topic>Chromatography</topic><topic>Clostridium thermocellum</topic><topic>Clostridium thermocellum - enzymology</topic><topic>dextrins</topic><topic>Dextrins - biosynthesis</topic><topic>Dextrins - chemistry</topic><topic>Dextrins - metabolism</topic><topic>enzyme activity</topic><topic>Fermentation</topic><topic>Fundamental and applied biological sciences. 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P</au><au>Lynd, L. R</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Biosynthesis of radiolabeled cellodextrins by the Clostridium thermocellum cellobiose and cellodextrin phosphorylases for measurement of intracellular sugars</atitle><jtitle>Applied microbiology and biotechnology</jtitle><addtitle>Appl Microbiol Biotechnol</addtitle><date>2006-03-01</date><risdate>2006</risdate><volume>70</volume><issue>1</issue><spage>123</spage><epage>129</epage><pages>123-129</pages><issn>0175-7598</issn><eissn>1432-0614</eissn><coden>AMBIDG</coden><abstract>The Clostridium thermocellum cellobiose and cellodextrin phosphorylases (glucosyl transferases) in the cell extract were used to synthesize radiolabeled cellodextrins with a degree of polymerization (DP=2-6) from nonradioactive glucose-1-phosphate and radioactive glucose. Chain lengths of synthesized cellodextrin were controlled by the absence or presence of dithiothreitol and by reaction conditions. All cellodextrins have the sole radioactive glucose unit located at the reducing ends. Mixed cellodextrins (G2-G6) were separated efficiently by size-exclusion chromatography or less efficiently by thin-layer chromatography. A new rapid sampling device was developed using disposable syringes containing an ultracold methanol-quenching buffer. It was simple, less costly, and especially convenient for anaerobic fermentation. After an impulse feed of radiolabeled cellobiose, the intracellular sugar levels were measured after a series of operations--sampling, extracting, concentrating, separating, and reading. Results showed that the largest amount of radioactivity was cellobiose with lesser amounts of glucose, cellotriose, and cellotetraose, and an average DP of intracellular cellodextrins was ca. 2.</abstract><cop>Berlin</cop><pub>Berlin/Heidelberg : Springer-Verlag</pub><pmid>16402169</pmid><doi>10.1007/s00253-005-0278-1</doi><tpages>7</tpages></addata></record> |
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| ispartof | Applied microbiology and biotechnology, 2006-03, Vol.70 (1), p.123-129 |
| issn | 0175-7598 1432-0614 |
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| source | ABI/INFORM global; Springer Link |
| subjects | Anaerobiosis Bacteriology Biological and medical sciences Biosynthesis Biotechnology Carbohydrate Conformation carbohydrate metabolism Carbon Radioisotopes cellobiose Cellulose - analogs & derivatives Cellulose - biosynthesis Cellulose - chemistry Cellulose - metabolism Chromatography Clostridium thermocellum Clostridium thermocellum - enzymology dextrins Dextrins - biosynthesis Dextrins - chemistry Dextrins - metabolism enzyme activity Fermentation Fundamental and applied biological sciences. Psychology Glucose Glucosyltransferases - metabolism hexosyltransferases Microbiology Radioactivity radiolabeling Sugar sugars Syringes Thin layer chromatography |
| title | Biosynthesis of radiolabeled cellodextrins by the Clostridium thermocellum cellobiose and cellodextrin phosphorylases for measurement of intracellular sugars |
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