The reaction of 3-diazonium-1,2,4-triazole with tryptophan at acid pH: A scintillation method for tryptophan determination in proteins
Unlike conventional aryl diazonium reagents which are usually reactive at alkaline pH, 3-diazonium-1,2,4-triazole (3-DT) has been found to react with a variety of proteins at acid pH. Using indole, phenol, and imidazole as models of aromatic amino acid side chains, the rate of 3-DT coupling to the i...
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| Veröffentlicht in: | Anal. Biochem.; (United States) 1979-11, Vol.99 (2), p.464-473 |
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| creator | DeTraglia, Michael C. Brand, John S. Tometsko, Andrew M. |
| description | Unlike conventional aryl diazonium reagents which are usually reactive at alkaline pH, 3-diazonium-1,2,4-triazole (3-DT) has been found to react with a variety of proteins at acid pH. Using indole, phenol, and imidazole as models of aromatic amino acid side chains, the rate of 3-DT coupling to the indole nucleus has been shown to increase exponentially from 2400
m
−1 min
−1 at pH 2.5. Coupling rates with phenol and imidazole show the conventional alkaline reactivity, with no evidence of reaction at acid pH. In parallel experiments using trypsin, 3-DT was found to cause loss in enzymatic activity at acidic as well as alkaline pH. Furthermore, amino acid analysis has demonstrated that the acidic reaction with trypsin resulted in the modification only of tryptophan. Based on the acidic specificity of 3-DT seen in these studies, a scintillation method for determining the tryptophan content of proteins has been developed. Using 3-diazonium-1,2,4-[5-
14C]triazole, accurate tryptophan contents for trypsin, chymotrypsin, lysozyme, pepsin, insulin, and fibrinogen have been measured. |
| doi_str_mv | 10.1016/S0003-2697(79)80034-2 |
| format | Article |
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m
−1 min
−1 at pH 6 to > 2400
m
−1 min
−1 at pH 2.5. Coupling rates with phenol and imidazole show the conventional alkaline reactivity, with no evidence of reaction at acid pH. In parallel experiments using trypsin, 3-DT was found to cause loss in enzymatic activity at acidic as well as alkaline pH. Furthermore, amino acid analysis has demonstrated that the acidic reaction with trypsin resulted in the modification only of tryptophan. Based on the acidic specificity of 3-DT seen in these studies, a scintillation method for determining the tryptophan content of proteins has been developed. Using 3-diazonium-1,2,4-[5-
14C]triazole, accurate tryptophan contents for trypsin, chymotrypsin, lysozyme, pepsin, insulin, and fibrinogen have been measured.</description><identifier>ISSN: 0003-2697</identifier><identifier>EISSN: 1096-0309</identifier><identifier>DOI: 10.1016/S0003-2697(79)80034-2</identifier><identifier>PMID: 574724</identifier><language>eng</language><publisher>United States: Elsevier Inc</publisher><subject>550201 - Biochemistry- Tracer Techniques ; 550501 - Metabolism- Tracer Techniques ; ABSORPTION SPECTRA ; AMINO ACIDS ; Amino Acids - analysis ; Animals ; AZOLES ; BASIC BIOLOGICAL SCIENCES ; BIOCHEMICAL REACTION KINETICS ; BLOOD COAGULATION FACTORS ; CARBON 14 COMPOUNDS ; CARBOXYLIC ACIDS ; Cattle ; CHEMICAL ANALYSIS ; CHYMOTRYPSIN ; COUNTING TECHNIQUES ; DATA ; DATA FORMS ; Diazonium Compounds ; ENZYMES ; EXPERIMENTAL DATA ; FIBRINOGEN ; GLOBULINS ; GLYCOSYL HYDROLASES ; HETEROCYCLIC ACIDS ; HETEROCYCLIC COMPOUNDS ; HORMONES ; HYDROLASES ; Indicators and Reagents ; INDOLES ; INFORMATION ; INSULIN ; ISOLATED VALUES ; ISOTOPE APPLICATIONS ; KINETICS ; LABELLED COMPOUNDS ; LYSOZYME ; NUMERICAL DATA ; ORGANIC ACIDS ; ORGANIC COMPOUNDS ; ORGANIC NITROGEN COMPOUNDS ; PEPSIN ; PEPTIDE HORMONES ; PEPTIDE HYDROLASES ; PH VALUE ; PROTEINS ; PYRROLES ; QUANTITATIVE CHEMICAL ANALYSIS ; REACTION KINETICS ; RESPONSE MODIFYING FACTORS ; SCINTILLATION COUNTING ; SPECTRA ; Spectrophotometry, Ultraviolet - methods ; TRACER TECHNIQUES ; Triazoles ; TRYPSIN ; TRYPTOPHAN ; Tryptophan - analysis</subject><ispartof>Anal. Biochem.; (United States), 1979-11, Vol.99 (2), p.464-473</ispartof><rights>1979 Academic Press, Inc.</rights><lds50>peer_reviewed</lds50><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c386t-49b545d4f19d0815b7350257a621cbb1616633f1f025e06cf618b29ea05a6bf63</citedby><cites>FETCH-LOGICAL-c386t-49b545d4f19d0815b7350257a621cbb1616633f1f025e06cf618b29ea05a6bf63</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktohtml>$$Uhttps://dx.doi.org/10.1016/S0003-2697(79)80034-2$$EHTML$$P50$$Gelsevier$$H</linktohtml><link.rule.ids>315,781,785,886,3551,27926,27927,45997</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/574724$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink><backlink>$$Uhttps://www.osti.gov/biblio/5410256$$D View this record in Osti.gov$$Hfree_for_read</backlink></links><search><creatorcontrib>DeTraglia, Michael C.</creatorcontrib><creatorcontrib>Brand, John S.</creatorcontrib><creatorcontrib>Tometsko, Andrew M.</creatorcontrib><creatorcontrib>Univ. of Rochester Medical Center, NY</creatorcontrib><title>The reaction of 3-diazonium-1,2,4-triazole with tryptophan at acid pH: A scintillation method for tryptophan determination in proteins</title><title>Anal. Biochem.; (United States)</title><addtitle>Anal Biochem</addtitle><description>Unlike conventional aryl diazonium reagents which are usually reactive at alkaline pH, 3-diazonium-1,2,4-triazole (3-DT) has been found to react with a variety of proteins at acid pH. Using indole, phenol, and imidazole as models of aromatic amino acid side chains, the rate of 3-DT coupling to the indole nucleus has been shown to increase exponentially from <200
m
−1 min
−1 at pH 6 to > 2400
m
−1 min
−1 at pH 2.5. Coupling rates with phenol and imidazole show the conventional alkaline reactivity, with no evidence of reaction at acid pH. In parallel experiments using trypsin, 3-DT was found to cause loss in enzymatic activity at acidic as well as alkaline pH. Furthermore, amino acid analysis has demonstrated that the acidic reaction with trypsin resulted in the modification only of tryptophan. Based on the acidic specificity of 3-DT seen in these studies, a scintillation method for determining the tryptophan content of proteins has been developed. Using 3-diazonium-1,2,4-[5-
14C]triazole, accurate tryptophan contents for trypsin, chymotrypsin, lysozyme, pepsin, insulin, and fibrinogen have been measured.</description><subject>550201 - Biochemistry- Tracer Techniques</subject><subject>550501 - Metabolism- Tracer Techniques</subject><subject>ABSORPTION SPECTRA</subject><subject>AMINO ACIDS</subject><subject>Amino Acids - analysis</subject><subject>Animals</subject><subject>AZOLES</subject><subject>BASIC BIOLOGICAL SCIENCES</subject><subject>BIOCHEMICAL REACTION KINETICS</subject><subject>BLOOD COAGULATION FACTORS</subject><subject>CARBON 14 COMPOUNDS</subject><subject>CARBOXYLIC ACIDS</subject><subject>Cattle</subject><subject>CHEMICAL ANALYSIS</subject><subject>CHYMOTRYPSIN</subject><subject>COUNTING TECHNIQUES</subject><subject>DATA</subject><subject>DATA FORMS</subject><subject>Diazonium Compounds</subject><subject>ENZYMES</subject><subject>EXPERIMENTAL DATA</subject><subject>FIBRINOGEN</subject><subject>GLOBULINS</subject><subject>GLYCOSYL HYDROLASES</subject><subject>HETEROCYCLIC ACIDS</subject><subject>HETEROCYCLIC COMPOUNDS</subject><subject>HORMONES</subject><subject>HYDROLASES</subject><subject>Indicators and Reagents</subject><subject>INDOLES</subject><subject>INFORMATION</subject><subject>INSULIN</subject><subject>ISOLATED VALUES</subject><subject>ISOTOPE APPLICATIONS</subject><subject>KINETICS</subject><subject>LABELLED COMPOUNDS</subject><subject>LYSOZYME</subject><subject>NUMERICAL DATA</subject><subject>ORGANIC ACIDS</subject><subject>ORGANIC COMPOUNDS</subject><subject>ORGANIC NITROGEN COMPOUNDS</subject><subject>PEPSIN</subject><subject>PEPTIDE HORMONES</subject><subject>PEPTIDE HYDROLASES</subject><subject>PH VALUE</subject><subject>PROTEINS</subject><subject>PYRROLES</subject><subject>QUANTITATIVE CHEMICAL ANALYSIS</subject><subject>REACTION KINETICS</subject><subject>RESPONSE MODIFYING FACTORS</subject><subject>SCINTILLATION COUNTING</subject><subject>SPECTRA</subject><subject>Spectrophotometry, Ultraviolet - methods</subject><subject>TRACER TECHNIQUES</subject><subject>Triazoles</subject><subject>TRYPSIN</subject><subject>TRYPTOPHAN</subject><subject>Tryptophan - analysis</subject><issn>0003-2697</issn><issn>1096-0309</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1979</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFkc-O1SAUxonx33X0DTQhszCaDAot0NaNmUzUMZnEheOaUDikmFuowNWMD-BzS28nE3euCOf8vnM--BB6wegbRpl8-5VS2pJGDt2rbnjd1wsnzT20Y3SQhLZ0uI92d8hj9CTn75QyxoV8hB6KjncN36E_1xPgBNoUHwOODrfEev07Bn-YCTtrzjgpaS3sAf_yZcIl3SwlLpMOWBesjbd4uXyHz3E2PhS_3-vjpBnKFC12Mf2rsFAgzT5sjA94SbGAD_kpeuD0PsOz2_MEffv44friklx9-fT54vyKmLaXhfBhFFxY7thgac_E2LWCNqLTsmFmHJlkUratY64WgUrjJOvHZgBNhZajk-0JOt3mxly8qpYLmMnEEMAUJTiruhV6uUHV3Y8D5KJmnw3UpwWIh6w63veyZysoNtCkmHMCp5bkZ51uFKNqzUgdM1JrAKob1DEj1VTd89sFh3EGe6faQqnt91sb6k_89JBWoxAMWJ9Wnzb6_yz4C2vjoO0</recordid><startdate>19791101</startdate><enddate>19791101</enddate><creator>DeTraglia, Michael C.</creator><creator>Brand, John S.</creator><creator>Tometsko, Andrew M.</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><scope>OTOTI</scope></search><sort><creationdate>19791101</creationdate><title>The reaction of 3-diazonium-1,2,4-triazole with tryptophan at acid pH: A scintillation method for tryptophan determination in proteins</title><author>DeTraglia, Michael C. ; Brand, John S. ; Tometsko, Andrew M.</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c386t-49b545d4f19d0815b7350257a621cbb1616633f1f025e06cf618b29ea05a6bf63</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1979</creationdate><topic>550201 - Biochemistry- Tracer Techniques</topic><topic>550501 - Metabolism- Tracer Techniques</topic><topic>ABSORPTION SPECTRA</topic><topic>AMINO ACIDS</topic><topic>Amino Acids - analysis</topic><topic>Animals</topic><topic>AZOLES</topic><topic>BASIC BIOLOGICAL SCIENCES</topic><topic>BIOCHEMICAL REACTION KINETICS</topic><topic>BLOOD COAGULATION FACTORS</topic><topic>CARBON 14 COMPOUNDS</topic><topic>CARBOXYLIC ACIDS</topic><topic>Cattle</topic><topic>CHEMICAL ANALYSIS</topic><topic>CHYMOTRYPSIN</topic><topic>COUNTING TECHNIQUES</topic><topic>DATA</topic><topic>DATA FORMS</topic><topic>Diazonium Compounds</topic><topic>ENZYMES</topic><topic>EXPERIMENTAL DATA</topic><topic>FIBRINOGEN</topic><topic>GLOBULINS</topic><topic>GLYCOSYL HYDROLASES</topic><topic>HETEROCYCLIC ACIDS</topic><topic>HETEROCYCLIC COMPOUNDS</topic><topic>HORMONES</topic><topic>HYDROLASES</topic><topic>Indicators and Reagents</topic><topic>INDOLES</topic><topic>INFORMATION</topic><topic>INSULIN</topic><topic>ISOLATED VALUES</topic><topic>ISOTOPE APPLICATIONS</topic><topic>KINETICS</topic><topic>LABELLED COMPOUNDS</topic><topic>LYSOZYME</topic><topic>NUMERICAL DATA</topic><topic>ORGANIC ACIDS</topic><topic>ORGANIC COMPOUNDS</topic><topic>ORGANIC NITROGEN COMPOUNDS</topic><topic>PEPSIN</topic><topic>PEPTIDE HORMONES</topic><topic>PEPTIDE HYDROLASES</topic><topic>PH VALUE</topic><topic>PROTEINS</topic><topic>PYRROLES</topic><topic>QUANTITATIVE CHEMICAL ANALYSIS</topic><topic>REACTION KINETICS</topic><topic>RESPONSE MODIFYING FACTORS</topic><topic>SCINTILLATION COUNTING</topic><topic>SPECTRA</topic><topic>Spectrophotometry, Ultraviolet - methods</topic><topic>TRACER TECHNIQUES</topic><topic>Triazoles</topic><topic>TRYPSIN</topic><topic>TRYPTOPHAN</topic><topic>Tryptophan - analysis</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>DeTraglia, Michael C.</creatorcontrib><creatorcontrib>Brand, John S.</creatorcontrib><creatorcontrib>Tometsko, Andrew M.</creatorcontrib><creatorcontrib>Univ. of Rochester Medical Center, NY</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><collection>OSTI.GOV</collection><jtitle>Anal. Biochem.; (United States)</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>DeTraglia, Michael C.</au><au>Brand, John S.</au><au>Tometsko, Andrew M.</au><aucorp>Univ. of Rochester Medical Center, NY</aucorp><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>The reaction of 3-diazonium-1,2,4-triazole with tryptophan at acid pH: A scintillation method for tryptophan determination in proteins</atitle><jtitle>Anal. Biochem.; (United States)</jtitle><addtitle>Anal Biochem</addtitle><date>1979-11-01</date><risdate>1979</risdate><volume>99</volume><issue>2</issue><spage>464</spage><epage>473</epage><pages>464-473</pages><issn>0003-2697</issn><eissn>1096-0309</eissn><abstract>Unlike conventional aryl diazonium reagents which are usually reactive at alkaline pH, 3-diazonium-1,2,4-triazole (3-DT) has been found to react with a variety of proteins at acid pH. Using indole, phenol, and imidazole as models of aromatic amino acid side chains, the rate of 3-DT coupling to the indole nucleus has been shown to increase exponentially from <200
m
−1 min
−1 at pH 6 to > 2400
m
−1 min
−1 at pH 2.5. Coupling rates with phenol and imidazole show the conventional alkaline reactivity, with no evidence of reaction at acid pH. In parallel experiments using trypsin, 3-DT was found to cause loss in enzymatic activity at acidic as well as alkaline pH. Furthermore, amino acid analysis has demonstrated that the acidic reaction with trypsin resulted in the modification only of tryptophan. Based on the acidic specificity of 3-DT seen in these studies, a scintillation method for determining the tryptophan content of proteins has been developed. Using 3-diazonium-1,2,4-[5-
14C]triazole, accurate tryptophan contents for trypsin, chymotrypsin, lysozyme, pepsin, insulin, and fibrinogen have been measured.</abstract><cop>United States</cop><pub>Elsevier Inc</pub><pmid>574724</pmid><doi>10.1016/S0003-2697(79)80034-2</doi><tpages>10</tpages></addata></record> |
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| ispartof | Anal. Biochem.; (United States), 1979-11, Vol.99 (2), p.464-473 |
| issn | 0003-2697 1096-0309 |
| language | eng |
| recordid | cdi_proquest_miscellaneous_74886816 |
| source | MEDLINE; Access via ScienceDirect (Elsevier) |
| subjects | 550201 - Biochemistry- Tracer Techniques 550501 - Metabolism- Tracer Techniques ABSORPTION SPECTRA AMINO ACIDS Amino Acids - analysis Animals AZOLES BASIC BIOLOGICAL SCIENCES BIOCHEMICAL REACTION KINETICS BLOOD COAGULATION FACTORS CARBON 14 COMPOUNDS CARBOXYLIC ACIDS Cattle CHEMICAL ANALYSIS CHYMOTRYPSIN COUNTING TECHNIQUES DATA DATA FORMS Diazonium Compounds ENZYMES EXPERIMENTAL DATA FIBRINOGEN GLOBULINS GLYCOSYL HYDROLASES HETEROCYCLIC ACIDS HETEROCYCLIC COMPOUNDS HORMONES HYDROLASES Indicators and Reagents INDOLES INFORMATION INSULIN ISOLATED VALUES ISOTOPE APPLICATIONS KINETICS LABELLED COMPOUNDS LYSOZYME NUMERICAL DATA ORGANIC ACIDS ORGANIC COMPOUNDS ORGANIC NITROGEN COMPOUNDS PEPSIN PEPTIDE HORMONES PEPTIDE HYDROLASES PH VALUE PROTEINS PYRROLES QUANTITATIVE CHEMICAL ANALYSIS REACTION KINETICS RESPONSE MODIFYING FACTORS SCINTILLATION COUNTING SPECTRA Spectrophotometry, Ultraviolet - methods TRACER TECHNIQUES Triazoles TRYPSIN TRYPTOPHAN Tryptophan - analysis |
| title | The reaction of 3-diazonium-1,2,4-triazole with tryptophan at acid pH: A scintillation method for tryptophan determination in proteins |
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