{"id":{"repo_id":"vt","oai_identifier":"oai:vtechworks.lib.vt.edu:10919/37625"},"canonical_url":"https://search.dev.ndltd.org/etd/vt/oai:vtechworks.lib.vt.edu:10919/37625","repository":{"repo_id":"vt","name":"Virginia Tech","base_url":"https://vtechworks.lib.vt.edu/oai/request"},"display":{"title":"Dual-specific protein phosphatases in the <i>Archaea</i>","abstract":"Three distinct families of PTPs, the conventional (cPTPs), low molecular weight (LMW PTPs), and Cdc25 PTPs, have converged upon a common catalytic mechanism and active site sequence, mainly, the phosphate-binding loop encompassing the PTP signature motif (H/V)<b>C</b>(X)₅<b>R</b>(S/T) and an essential Asp residue on a surface loop. There is little sequence similarity among the three families of phosphatases. All known LMW PTP remove phosphoryl groups esterified to the hydroxyl amino acid: tyrosine, whereas all members of the Cdc25 family are dual-specificity protein phosphatases that dephosphorylate all the hydroxyl amino acids: tyrosine, serine and threonine. The cPTP family primarily functions as tyrosine phosphatases, but it also includes dual-specific members. ORFs encoding potential cPTPs have been identified in five archaeal species: <i>Methanobacterium thermoautotrophicum</i>, <i>Methanococcus jannaschii</i>, <i>Thermococcus kodakaraensis</i>, <i>Pyrococcus horikoshii</i>, and <i>S. solfataricus</i>. Only one has been partially characterized, <i>Tk</i>-PTP from <i>T. kodakaraensis</i>. Hence, our current body of knowledge concerning the functional properties and physiological roles of these enzymes remains fragmented. The genome of <i>S. solfataricus</i> encodes a single conventional protein tyrosine phosphatase, SsoPTP. SsoPTP is the smallest known archaeal PTP (18.3 kDa) with a primary amino acid sequence that conforms to the cPTP protein tyrosine phosphatase paradigm, HCX₅R(S/T). Relatively little is known about its mode of action \" whether it follows the conventional PTP mechanism or employs a different route for catalysis \" or its physiological role. ORF <i>sso2453</i> from the genome of <i>Sulfolobus solfataricus</i>, encoding a protein tyrosine phosphatase, was cloned and its recombinant protein product, SsoPTP, was expressed in <i>E. coli</i> and purified by immobilized metal affinity chromatography. SsoPTP displayed the ability to dephosphorylate protein-bound phosphotyrosine as well as protein-bound phosphoserine/phosphothreonine. SsoPTP hydrolyzed both isomers of naphthyl phosphate, an indication of dual specificity. The four conserved residues within the presumed active site sequence: Asp⁶⁹, His⁹⁵, and Arg¹⁰², and the invariant Gln¹³⁹ residue were essential for catalysis, as it was predicted for the established members of the PTP family in both bacteria and eukaryotes. A substrate trapping protein variant, SsoPTP-C96S/D69A, was constructed to isolate possible SsoPTP substrates present in <i>S. solfataricus</i> cell lysates. Several potential substrates were isolated and identified by mass spectroscopy.","abstract_html":"Three distinct families of PTPs, the conventional (cPTPs), low molecular weight (LMW PTPs), and Cdc25 PTPs, have converged upon a common catalytic mechanism and active site sequence, mainly, the phosphate-binding loop encompassing the PTP signature motif (H/V)&lt;b&gt;C&lt;/b&gt;(X)₅&lt;b&gt;R&lt;/b&gt;(S/T) and an essential Asp residue on a surface loop. There is little sequence similarity among the three families of phosphatases. All known LMW PTP remove phosphoryl groups esterified to the hydroxyl amino acid: tyrosine, whereas all members of the Cdc25 family are dual-specificity protein phosphatases that dephosphorylate all the hydroxyl amino acids: tyrosine, serine and threonine. The cPTP family primarily functions as tyrosine phosphatases, but it also includes dual-specific members. ORFs encoding potential cPTPs have been identified in five archaeal species: &lt;i&gt;Methanobacterium thermoautotrophicum&lt;/i&gt;, &lt;i&gt;Methanococcus jannaschii&lt;/i&gt;, &lt;i&gt;Thermococcus kodakaraensis&lt;/i&gt;, &lt;i&gt;Pyrococcus horikoshii&lt;/i&gt;, and &lt;i&gt;S. solfataricus&lt;/i&gt;. Only one has been partially characterized, &lt;i&gt;Tk&lt;/i&gt;-PTP from &lt;i&gt;T. kodakaraensis&lt;/i&gt;. Hence, our current body of knowledge concerning the functional properties and physiological roles of these enzymes remains fragmented. The genome of &lt;i&gt;S. solfataricus&lt;/i&gt; encodes a single conventional protein tyrosine phosphatase, SsoPTP. SsoPTP is the smallest known archaeal PTP (18.3 kDa) with a primary amino acid sequence that conforms to the cPTP protein tyrosine phosphatase paradigm, HCX₅R(S/T). Relatively little is known about its mode of action &quot; whether it follows the conventional PTP mechanism or employs a different route for catalysis &quot; or its physiological role. ORF &lt;i&gt;sso2453&lt;/i&gt; from the genome of &lt;i&gt;Sulfolobus solfataricus&lt;/i&gt;, encoding a protein tyrosine phosphatase, was cloned and its recombinant protein product, SsoPTP, was expressed in &lt;i&gt;E. coli&lt;/i&gt; and purified by immobilized metal affinity chromatography. SsoPTP displayed the ability to dephosphorylate protein-bound phosphotyrosine as well as protein-bound phosphoserine/phosphothreonine. SsoPTP hydrolyzed both isomers of naphthyl phosphate, an indication of dual specificity. The four conserved residues within the presumed active site sequence: Asp⁶⁹, His⁹⁵, and Arg¹⁰², and the invariant Gln¹³⁹ residue were essential for catalysis, as it was predicted for the established members of the PTP family in both bacteria and eukaryotes. A substrate trapping protein variant, SsoPTP-C96S/D69A, was constructed to isolate possible SsoPTP substrates present in &lt;i&gt;S. solfataricus&lt;/i&gt; cell lysates. Several potential substrates were isolated and identified by mass spectroscopy.","abstract_has_math":false,"creators":["Dahche, Hanan Mohamad"],"institution":"Virginia Tech","degree_name":"Ph. D.","degree_level":"doctoral","degree_discipline":"Biochemistry","degree_department":"Biochemistry","school":null,"contributors":[],"advisors":[],"committee_chairs":["Kennelly, Peter J."],"committee_members":["Li, Jianyong","Helm, Richard F.","Bevan, David R."],"year":2010,"date_issued":"2010-04-02","date_published":"2010-04-02","updated_at":"2026-07-22T22:19:56Z","subjects":["dual-specific phosphatase","Archaea","protein tyrosine phosphatase","protein phosphorylation"],"languages":[],"rights":["In Copyright"],"rights_urls":["http://rightsstatements.org/vocab/InC/1.0/"],"identifier_entries":[{"key":"dc:identifier.other","label":"Dc Identifier Other","values":["etd-04142010-151033"],"render_values":[{"text":"etd-04142010-151033","href":null,"code":true}]}]},"links":{"outbound_url":"http://hdl.handle.net/10919/37625","outbound_label":"Handle","outbound_source":"dc:identifier.uri"},"metadata_groups":[{"id":"people","label":"People","entries":[{"key":"dc:contributor.committeechair","label":"Committee Chair","values":["Kennelly, Peter J."]},{"key":"dc:contributor.committeemember","label":"Committee Member","values":["Li, Jianyong","Helm, Richard F.","Bevan, David R."]},{"key":"dc:contributor.department","label":"Department","values":["Biochemistry"]},{"key":"dc:creator","label":"Author","values":["Dahche, Hanan Mohamad"]}]},{"id":"academic_context","label":"Academic Context","entries":[{"key":"dc:date.accessioned","label":"Dc Date Accessioned","values":["2014-03-14T21:10:30Z"]},{"key":"dc:date.available","label":"Dc Date Available","values":["2014-03-14T21:10:30Z","2010-05-03"]},{"key":"dc:date.issued","label":"Date","values":["2010-04-02"]},{"key":"dc:publisher","label":"Institution","values":["Virginia Tech"]},{"key":"dc:type","label":"Dc Type","values":["Dissertation"]},{"key":"thesis:degree_discipline","label":"Discipline","values":["Biochemistry"]},{"key":"thesis:degree_level","label":"Degree Level","values":["doctoral"]},{"key":"thesis:degree_name","label":"Degree Name","values":["Ph. D."]},{"key":"thesis:institution_name","label":"Thesis Institution Name","values":["Virginia Polytechnic Institute and State University"]}]},{"id":"subjects_keywords","label":"Subjects and Keywords","entries":[{"key":"dc:subject","label":"Dc Subject","values":["dual-specific phosphatase","Archaea","protein tyrosine phosphatase","protein phosphorylation"]}]},{"id":"language_rights","label":"Language and Rights","entries":[{"key":"dc:rights","label":"Dc Rights","values":["In Copyright"]},{"key":"dc:rights.uri","label":"Rights URI","values":["http://rightsstatements.org/vocab/InC/1.0/"]}]},{"id":"identifiers","label":"Identifiers","entries":[{"key":"dc:identifier.other","label":"Dc Identifier Other","values":["etd-04142010-151033"]},{"key":"dc:identifier.uri","label":"Identifier URI","values":["http://hdl.handle.net/10919/37625"]}]},{"id":"additional","label":"Additional Metadata","entries":[{"key":"dc:description.abstract","label":"Abstract","values":["Three distinct families of PTPs, the conventional (cPTPs), low molecular weight (LMW PTPs), and Cdc25 PTPs, have converged upon a common catalytic mechanism and active site sequence, mainly, the phosphate-binding loop encompassing the PTP signature motif (H/V)<b>C</b>(X)₅<b>R</b>(S/T) and an essential Asp residue on a surface loop. There is little sequence similarity among the three families of phosphatases. All known LMW PTP remove phosphoryl groups esterified to the hydroxyl amino acid: tyrosine, whereas all members of the Cdc25 family are dual-specificity protein phosphatases that dephosphorylate all the hydroxyl amino acids: tyrosine, serine and threonine. The cPTP family primarily functions as tyrosine phosphatases, but it also includes dual-specific members. ORFs encoding potential cPTPs have been identified in five archaeal species: <i>Methanobacterium thermoautotrophicum</i>, <i>Methanococcus jannaschii</i>, <i>Thermococcus kodakaraensis</i>, <i>Pyrococcus horikoshii</i>, and <i>S. solfataricus</i>. Only one has been partially characterized, <i>Tk</i>-PTP from <i>T. kodakaraensis</i>. Hence, our current body of knowledge concerning the functional properties and physiological roles of these enzymes remains fragmented. The genome of <i>S. solfataricus</i> encodes a single conventional protein tyrosine phosphatase, SsoPTP. SsoPTP is the smallest known archaeal PTP (18.3 kDa) with a primary amino acid sequence that conforms to the cPTP protein tyrosine phosphatase paradigm, HCX₅R(S/T). Relatively little is known about its mode of action \" whether it follows the conventional PTP mechanism or employs a different route for catalysis \" or its physiological role. ORF <i>sso2453</i> from the genome of <i>Sulfolobus solfataricus</i>, encoding a protein tyrosine phosphatase, was cloned and its recombinant protein product, SsoPTP, was expressed in <i>E. coli</i> and purified by immobilized metal affinity chromatography. SsoPTP displayed the ability to dephosphorylate protein-bound phosphotyrosine as well as protein-bound phosphoserine/phosphothreonine. SsoPTP hydrolyzed both isomers of naphthyl phosphate, an indication of dual specificity. The four conserved residues within the presumed active site sequence: Asp⁶⁹, His⁹⁵, and Arg¹⁰², and the invariant Gln¹³⁹ residue were essential for catalysis, as it was predicted for the established members of the PTP family in both bacteria and eukaryotes. A substrate trapping protein variant, SsoPTP-C96S/D69A, was constructed to isolate possible SsoPTP substrates present in <i>S. solfataricus</i> cell lysates. Several potential substrates were isolated and identified by mass spectroscopy."]},{"key":"dc:description.degree","label":"Dc Description Degree","values":["Ph. D."]},{"key":"dc:title","label":"Title","values":["Dual-specific protein phosphatases in the <i>Archaea</i>"]}]}],"canonical_facts":{"dc:contributor.committeechair":["Kennelly, Peter J."],"dc:contributor.committeemember":["Li, Jianyong","Helm, Richard F.","Bevan, David R."],"dc:contributor.department":["Biochemistry"],"dc:creator":["Dahche, Hanan Mohamad"],"dc:date.accessioned":["2014-03-14T21:10:30Z"],"dc:date.available":["2014-03-14T21:10:30Z","2010-05-03"],"dc:date.issued":["2010-04-02"],"dc:description.abstract":["Three distinct families of PTPs, the conventional (cPTPs), low molecular weight (LMW PTPs), and Cdc25 PTPs, have converged upon a common catalytic mechanism and active site sequence, mainly, the phosphate-binding loop encompassing the PTP signature motif (H/V)<b>C</b>(X)₅<b>R</b>(S/T) and an essential Asp residue on a surface loop. There is little sequence similarity among the three families of phosphatases. All known LMW PTP remove phosphoryl groups esterified to the hydroxyl amino acid: tyrosine, whereas all members of the Cdc25 family are dual-specificity protein phosphatases that dephosphorylate all the hydroxyl amino acids: tyrosine, serine and threonine. The cPTP family primarily functions as tyrosine phosphatases, but it also includes dual-specific members. ORFs encoding potential cPTPs have been identified in five archaeal species: <i>Methanobacterium thermoautotrophicum</i>, <i>Methanococcus jannaschii</i>, <i>Thermococcus kodakaraensis</i>, <i>Pyrococcus horikoshii</i>, and <i>S. solfataricus</i>. Only one has been partially characterized, <i>Tk</i>-PTP from <i>T. kodakaraensis</i>. Hence, our current body of knowledge concerning the functional properties and physiological roles of these enzymes remains fragmented. The genome of <i>S. solfataricus</i> encodes a single conventional protein tyrosine phosphatase, SsoPTP. SsoPTP is the smallest known archaeal PTP (18.3 kDa) with a primary amino acid sequence that conforms to the cPTP protein tyrosine phosphatase paradigm, HCX₅R(S/T). Relatively little is known about its mode of action \" whether it follows the conventional PTP mechanism or employs a different route for catalysis \" or its physiological role. ORF <i>sso2453</i> from the genome of <i>Sulfolobus solfataricus</i>, encoding a protein tyrosine phosphatase, was cloned and its recombinant protein product, SsoPTP, was expressed in <i>E. coli</i> and purified by immobilized metal affinity chromatography. SsoPTP displayed the ability to dephosphorylate protein-bound phosphotyrosine as well as protein-bound phosphoserine/phosphothreonine. SsoPTP hydrolyzed both isomers of naphthyl phosphate, an indication of dual specificity. The four conserved residues within the presumed active site sequence: Asp⁶⁹, His⁹⁵, and Arg¹⁰², and the invariant Gln¹³⁹ residue were essential for catalysis, as it was predicted for the established members of the PTP family in both bacteria and eukaryotes. A substrate trapping protein variant, SsoPTP-C96S/D69A, was constructed to isolate possible SsoPTP substrates present in <i>S. solfataricus</i> cell lysates. Several potential substrates were isolated and identified by mass spectroscopy."],"dc:description.degree":["Ph. D."],"dc:identifier.other":["etd-04142010-151033"],"dc:identifier.uri":["http://hdl.handle.net/10919/37625"],"dc:publisher":["Virginia Tech"],"dc:rights":["In Copyright"],"dc:rights.uri":["http://rightsstatements.org/vocab/InC/1.0/"],"dc:subject":["dual-specific phosphatase","Archaea","protein tyrosine phosphatase","protein phosphorylation"],"dc:title":["Dual-specific protein phosphatases in the <i>Archaea</i>"],"dc:type":["Dissertation"],"thesis:degree_discipline":["Biochemistry"],"thesis:degree_level":["doctoral"],"thesis:degree_name":["Ph. D."],"thesis:institution_name":["Virginia Polytechnic Institute and State University"]},"updated_at":"2026-07-22T22:19:56Z"}