Biochemical characterization and mode of action of a thermostable endoglucanase purified from Thermoascus aurantiacus
A major extracellular endoglucanase purified to homogeneity from Thermoascus aurantiacus had a M r of 34 kDa and a p I of 3.7 and was optimally active at 70–80 °C and pH 4.0–4.4. It was stable at pH 2.8–6.8 at 50 °C for 48 h and maintained its secondary structure and folded conformation up to 70 °C...
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| Veröffentlicht in: | Archives of biochemistry and biophysics 2002-08, Vol.404 (2), p.243-253 |
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| container_title | Archives of biochemistry and biophysics |
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| creator | Parry, N.J Beever, D.E Owen, E Nerinckx, W Claeyssens, M Van Beeumen, J Bhat, M.K |
| description | A major extracellular endoglucanase purified to homogeneity from
Thermoascus aurantiacus had a
M
r of 34
kDa and a p
I of 3.7 and was optimally active at 70–80
°C and pH 4.0–4.4. It was stable at pH 2.8–6.8 at 50
°C for 48
h and maintained its secondary structure and folded conformation up to 70
°C at pH 5.0 and 2.8, respectively. A 33-amino acid sequence at the N terminus showed considerable homology with 14 microbial endoglucanases having highly conserved 8 amino acids (positions 10–17) and Gly, Pro, Gly, and Pro at positions 8, 22, 23, and 32, respectively. The enzyme is rich in Asp (15%) and Glu (10%) with a carbohydrate content of 2.7%. Polyclonal antibodies of endoglucanase cross-reacted with their own antigen and with other purified cellulases from
T. aurantiacus. The endoglucanase was specific for polymeric substrates with highest activity toward carboxymethyl cellulose followed by barley β-glucan and lichenan. It preferentially cleaved the internal glycosidic bonds of Glc
n
and MeUmbGlc
n
and possessed an extended substrate-binding site with five subsites. The data indicate that the endoglucanase from
T. aurantiacus is a member of glycoside hydrolase family 5. |
| doi_str_mv | 10.1016/S0003-9861(02)00301-6 |
| format | Article |
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Thermoascus aurantiacus had a
M
r of 34
kDa and a p
I of 3.7 and was optimally active at 70–80
°C and pH 4.0–4.4. It was stable at pH 2.8–6.8 at 50
°C for 48
h and maintained its secondary structure and folded conformation up to 70
°C at pH 5.0 and 2.8, respectively. A 33-amino acid sequence at the N terminus showed considerable homology with 14 microbial endoglucanases having highly conserved 8 amino acids (positions 10–17) and Gly, Pro, Gly, and Pro at positions 8, 22, 23, and 32, respectively. The enzyme is rich in Asp (15%) and Glu (10%) with a carbohydrate content of 2.7%. Polyclonal antibodies of endoglucanase cross-reacted with their own antigen and with other purified cellulases from
T. aurantiacus. The endoglucanase was specific for polymeric substrates with highest activity toward carboxymethyl cellulose followed by barley β-glucan and lichenan. It preferentially cleaved the internal glycosidic bonds of Glc
n
and MeUmbGlc
n
and possessed an extended substrate-binding site with five subsites. The data indicate that the endoglucanase from
T. aurantiacus is a member of glycoside hydrolase family 5.</description><identifier>ISSN: 0003-9861</identifier><identifier>EISSN: 1096-0384</identifier><identifier>DOI: 10.1016/S0003-9861(02)00301-6</identifier><identifier>PMID: 12147262</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>Ascomycota - enzymology ; Calorimetry, Differential Scanning ; Carboxymethylcellulose Sodium - chemistry ; Cellulase - chemistry ; Cellulase - isolation & purification ; Chromatography, High Pressure Liquid ; Endoglucanase ; Enzyme Activation - physiology ; Enzyme Stability - physiology ; Extracellular Space - enzymology ; Glucans - chemistry ; Glycoside hydrolase family 5 ; Hydrogen-Ion Concentration ; Mode of action ; Molecular Sequence Data ; N-terminal sequence ; Protein Conformation ; Protein Folding ; Protein Structure, Secondary ; Sequence Analysis, Protein ; Sequence Homology, Amino Acid ; Stability ; Substrate specificity ; Substrate Specificity - physiology ; T. aurantiacus ; Temperature ; Thermodynamics</subject><ispartof>Archives of biochemistry and biophysics, 2002-08, Vol.404 (2), p.243-253</ispartof><rights>2002 Elsevier Science (USA)</rights><rights>Copyright 2002 Elsevier Science (USA).</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c408t-9623c457ddaa51b0672e7950fd39c917f0bc2811be4c342905b2619b820b28163</citedby><cites>FETCH-LOGICAL-c408t-9623c457ddaa51b0672e7950fd39c917f0bc2811be4c342905b2619b820b28163</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://www.sciencedirect.com/science/article/pii/S0003986102003016$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>314,776,780,3537,27901,27902,65306</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/12147262$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Parry, N.J</creatorcontrib><creatorcontrib>Beever, D.E</creatorcontrib><creatorcontrib>Owen, E</creatorcontrib><creatorcontrib>Nerinckx, W</creatorcontrib><creatorcontrib>Claeyssens, M</creatorcontrib><creatorcontrib>Van Beeumen, J</creatorcontrib><creatorcontrib>Bhat, M.K</creatorcontrib><title>Biochemical characterization and mode of action of a thermostable endoglucanase purified from Thermoascus aurantiacus</title><title>Archives of biochemistry and biophysics</title><addtitle>Arch Biochem Biophys</addtitle><description>A major extracellular endoglucanase purified to homogeneity from
Thermoascus aurantiacus had a
M
r of 34
kDa and a p
I of 3.7 and was optimally active at 70–80
°C and pH 4.0–4.4. It was stable at pH 2.8–6.8 at 50
°C for 48
h and maintained its secondary structure and folded conformation up to 70
°C at pH 5.0 and 2.8, respectively. A 33-amino acid sequence at the N terminus showed considerable homology with 14 microbial endoglucanases having highly conserved 8 amino acids (positions 10–17) and Gly, Pro, Gly, and Pro at positions 8, 22, 23, and 32, respectively. The enzyme is rich in Asp (15%) and Glu (10%) with a carbohydrate content of 2.7%. Polyclonal antibodies of endoglucanase cross-reacted with their own antigen and with other purified cellulases from
T. aurantiacus. The endoglucanase was specific for polymeric substrates with highest activity toward carboxymethyl cellulose followed by barley β-glucan and lichenan. It preferentially cleaved the internal glycosidic bonds of Glc
n
and MeUmbGlc
n
and possessed an extended substrate-binding site with five subsites. The data indicate that the endoglucanase from
T. aurantiacus is a member of glycoside hydrolase family 5.</description><subject>Ascomycota - enzymology</subject><subject>Calorimetry, Differential Scanning</subject><subject>Carboxymethylcellulose Sodium - chemistry</subject><subject>Cellulase - chemistry</subject><subject>Cellulase - isolation & purification</subject><subject>Chromatography, High Pressure Liquid</subject><subject>Endoglucanase</subject><subject>Enzyme Activation - physiology</subject><subject>Enzyme Stability - physiology</subject><subject>Extracellular Space - enzymology</subject><subject>Glucans - chemistry</subject><subject>Glycoside hydrolase family 5</subject><subject>Hydrogen-Ion Concentration</subject><subject>Mode of action</subject><subject>Molecular Sequence Data</subject><subject>N-terminal sequence</subject><subject>Protein Conformation</subject><subject>Protein Folding</subject><subject>Protein Structure, Secondary</subject><subject>Sequence Analysis, Protein</subject><subject>Sequence Homology, Amino Acid</subject><subject>Stability</subject><subject>Substrate specificity</subject><subject>Substrate Specificity - physiology</subject><subject>T. aurantiacus</subject><subject>Temperature</subject><subject>Thermodynamics</subject><issn>0003-9861</issn><issn>1096-0384</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2002</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkE1v1DAQhi0EotvCTwD5hOghMOMkTnxCpeJLqsSBcrYm9oQ1SuLFTpDg15PsruDIya_Gj_1qHiGeIbxCQP36CwCUhWk1vgR1vWbAQj8QOwSjCyjb6qHY_UUuxGXO3wEQK60eiwtUWDVKq51Y3obo9jwGR4N0e0rkZk7hN80hTpImL8foWcZerhfbaEty3nMaY56pG1jy5OO3YXE0UWZ5WFLoA3vZpzjK-yNI2S1Z0pJomgOt-Yl41NOQ-en5vBJf37-7v_1Y3H3-8On25q5wFbRzYbQqXVU33hPV2IFuFDemht6XxhlseuicahE7rlxZKQN1pzSarlXQrXNdXokXp38PKf5YOM92DNnxMNDEccm2QaPLWlUrWJ9Al2LOiXt7SGGk9Msi2M23Pfq2m0wLyh59263g-blg6Ub2_16dBa_AmxPA65o_AyebXeDJsQ-J3Wx9DP-p-ANSgZDY</recordid><startdate>20020815</startdate><enddate>20020815</enddate><creator>Parry, N.J</creator><creator>Beever, D.E</creator><creator>Owen, E</creator><creator>Nerinckx, W</creator><creator>Claeyssens, M</creator><creator>Van Beeumen, J</creator><creator>Bhat, M.K</creator><general>Elsevier Inc</general><scope>CGR</scope><scope>CUY</scope><scope>CVF</scope><scope>ECM</scope><scope>EIF</scope><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7X8</scope></search><sort><creationdate>20020815</creationdate><title>Biochemical characterization and mode of action of a thermostable endoglucanase purified from Thermoascus aurantiacus</title><author>Parry, N.J ; Beever, D.E ; Owen, E ; Nerinckx, W ; Claeyssens, M ; Van Beeumen, J ; Bhat, M.K</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c408t-9623c457ddaa51b0672e7950fd39c917f0bc2811be4c342905b2619b820b28163</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2002</creationdate><topic>Ascomycota - enzymology</topic><topic>Calorimetry, Differential Scanning</topic><topic>Carboxymethylcellulose Sodium - chemistry</topic><topic>Cellulase - chemistry</topic><topic>Cellulase - isolation & purification</topic><topic>Chromatography, High Pressure Liquid</topic><topic>Endoglucanase</topic><topic>Enzyme Activation - physiology</topic><topic>Enzyme Stability - physiology</topic><topic>Extracellular Space - enzymology</topic><topic>Glucans - chemistry</topic><topic>Glycoside hydrolase family 5</topic><topic>Hydrogen-Ion Concentration</topic><topic>Mode of action</topic><topic>Molecular Sequence Data</topic><topic>N-terminal sequence</topic><topic>Protein Conformation</topic><topic>Protein Folding</topic><topic>Protein Structure, Secondary</topic><topic>Sequence Analysis, Protein</topic><topic>Sequence Homology, Amino Acid</topic><topic>Stability</topic><topic>Substrate specificity</topic><topic>Substrate Specificity - physiology</topic><topic>T. aurantiacus</topic><topic>Temperature</topic><topic>Thermodynamics</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Parry, N.J</creatorcontrib><creatorcontrib>Beever, D.E</creatorcontrib><creatorcontrib>Owen, E</creatorcontrib><creatorcontrib>Nerinckx, W</creatorcontrib><creatorcontrib>Claeyssens, M</creatorcontrib><creatorcontrib>Van Beeumen, J</creatorcontrib><creatorcontrib>Bhat, M.K</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>MEDLINE - Academic</collection><jtitle>Archives of biochemistry and biophysics</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Parry, N.J</au><au>Beever, D.E</au><au>Owen, E</au><au>Nerinckx, W</au><au>Claeyssens, M</au><au>Van Beeumen, J</au><au>Bhat, M.K</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Biochemical characterization and mode of action of a thermostable endoglucanase purified from Thermoascus aurantiacus</atitle><jtitle>Archives of biochemistry and biophysics</jtitle><addtitle>Arch Biochem Biophys</addtitle><date>2002-08-15</date><risdate>2002</risdate><volume>404</volume><issue>2</issue><spage>243</spage><epage>253</epage><pages>243-253</pages><issn>0003-9861</issn><eissn>1096-0384</eissn><abstract>A major extracellular endoglucanase purified to homogeneity from
Thermoascus aurantiacus had a
M
r of 34
kDa and a p
I of 3.7 and was optimally active at 70–80
°C and pH 4.0–4.4. It was stable at pH 2.8–6.8 at 50
°C for 48
h and maintained its secondary structure and folded conformation up to 70
°C at pH 5.0 and 2.8, respectively. A 33-amino acid sequence at the N terminus showed considerable homology with 14 microbial endoglucanases having highly conserved 8 amino acids (positions 10–17) and Gly, Pro, Gly, and Pro at positions 8, 22, 23, and 32, respectively. The enzyme is rich in Asp (15%) and Glu (10%) with a carbohydrate content of 2.7%. Polyclonal antibodies of endoglucanase cross-reacted with their own antigen and with other purified cellulases from
T. aurantiacus. The endoglucanase was specific for polymeric substrates with highest activity toward carboxymethyl cellulose followed by barley β-glucan and lichenan. It preferentially cleaved the internal glycosidic bonds of Glc
n
and MeUmbGlc
n
and possessed an extended substrate-binding site with five subsites. The data indicate that the endoglucanase from
T. aurantiacus is a member of glycoside hydrolase family 5.</abstract><cop>United States</cop><pub>Elsevier Inc</pub><pmid>12147262</pmid><doi>10.1016/S0003-9861(02)00301-6</doi><tpages>11</tpages><oa>free_for_read</oa></addata></record> |
| fulltext | fulltext |
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| ispartof | Archives of biochemistry and biophysics, 2002-08, Vol.404 (2), p.243-253 |
| issn | 0003-9861 1096-0384 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_71963524 |
| source | MEDLINE; Elsevier ScienceDirect Journals |
| subjects | Ascomycota - enzymology Calorimetry, Differential Scanning Carboxymethylcellulose Sodium - chemistry Cellulase - chemistry Cellulase - isolation & purification Chromatography, High Pressure Liquid Endoglucanase Enzyme Activation - physiology Enzyme Stability - physiology Extracellular Space - enzymology Glucans - chemistry Glycoside hydrolase family 5 Hydrogen-Ion Concentration Mode of action Molecular Sequence Data N-terminal sequence Protein Conformation Protein Folding Protein Structure, Secondary Sequence Analysis, Protein Sequence Homology, Amino Acid Stability Substrate specificity Substrate Specificity - physiology T. aurantiacus Temperature Thermodynamics |
| title | Biochemical characterization and mode of action of a thermostable endoglucanase purified from Thermoascus aurantiacus |
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