The Pichia pastoris PAS4 gene encodes a ubiquitin-conjugating enzyme required for peroxisome assembly
We report here the cloning and initial characterization of PAS4, a gene required for peroxisome assembly in the yeast Pichia pastoris. The PAS4 gene encodes a 24-kDa protein (Pas4p) that is located on the cytoplasmic surface of peroxisomes and is induced during peroxisome proliferation. Analysis of...
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| Veröffentlicht in: | The Journal of biological chemistry 1994-08, Vol.269 (34), p.21835-21844 |
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| container_title | The Journal of biological chemistry |
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| creator | Crane, D I Kalish, J E Gould, S J |
| description | We report here the cloning and initial characterization of PAS4, a gene required for peroxisome assembly in the yeast Pichia
pastoris. The PAS4 gene encodes a 24-kDa protein (Pas4p) that is located on the cytoplasmic surface of peroxisomes and is
induced during peroxisome proliferation. Analysis of the Pas4p sequence revealed a high degree of similarity to ubiquitin-conjugating
enzymes, particularly in the region surrounding the putative active-site cysteine residue with which ubiquitin forms a thioester
bond. As expected for a ubiquitin-conjugating enzyme, substitution of alanine or serine for the conserved active-site cysteine
residue abolished PAS4 function. In addition, a small amount of a 32 kDa form of Pas4p (the predicted size of a Pas4p-ubiquitin
conjugate) was detected both in vivo and in vitro. This species was eliminated by reducing agents and was not detected in
the cysteine to alanine substitution mutant, suggesting that it is a Pas4p-ubiquitin conjugate. Using a yeast strain that
overexpresses a Myc-ubiquitin fusion protein, we demonstrate directly that this conjugate contains ubiquitin. We conclude
from these observations that PAS4 is a member of the ubiquitin-conjugating enzyme gene family and that one or more ubiquitination
reactions are required for peroxisome assembly. |
| doi_str_mv | 10.1016/s0021-9258(17)31879-3 |
| format | Article |
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pastoris. The PAS4 gene encodes a 24-kDa protein (Pas4p) that is located on the cytoplasmic surface of peroxisomes and is
induced during peroxisome proliferation. Analysis of the Pas4p sequence revealed a high degree of similarity to ubiquitin-conjugating
enzymes, particularly in the region surrounding the putative active-site cysteine residue with which ubiquitin forms a thioester
bond. As expected for a ubiquitin-conjugating enzyme, substitution of alanine or serine for the conserved active-site cysteine
residue abolished PAS4 function. In addition, a small amount of a 32 kDa form of Pas4p (the predicted size of a Pas4p-ubiquitin
conjugate) was detected both in vivo and in vitro. This species was eliminated by reducing agents and was not detected in
the cysteine to alanine substitution mutant, suggesting that it is a Pas4p-ubiquitin conjugate. Using a yeast strain that
overexpresses a Myc-ubiquitin fusion protein, we demonstrate directly that this conjugate contains ubiquitin. We conclude
from these observations that PAS4 is a member of the ubiquitin-conjugating enzyme gene family and that one or more ubiquitination
reactions are required for peroxisome assembly.</description><identifier>ISSN: 0021-9258</identifier><identifier>EISSN: 1083-351X</identifier><identifier>DOI: 10.1016/s0021-9258(17)31879-3</identifier><identifier>PMID: 8063827</identifier><language>eng</language><publisher>United States: American Society for Biochemistry and Molecular Biology</publisher><subject>Amino Acid Sequence ; Base Sequence ; Cell Compartmentation ; Enzyme Induction ; Fungal Proteins ; Genes, Fungal - genetics ; Genes, myc - genetics ; Genetic Complementation Test ; Intracellular Membranes - chemistry ; Ligases - analysis ; Ligases - genetics ; Ligases - metabolism ; Microbodies - chemistry ; Microbodies - physiology ; Molecular Sequence Data ; Mutation ; Pichia - enzymology ; Pichia - genetics ; Pichia pastoris ; Recombinant Fusion Proteins - metabolism ; Sequence Homology, Amino Acid ; Ubiquitin-Conjugating Enzymes ; Ubiquitins - metabolism</subject><ispartof>The Journal of biological chemistry, 1994-08, Vol.269 (34), p.21835-21844</ispartof><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c477t-6e82d5da697c38f62d141d3f1fd75a7df66de683bc5d7d34046df0742be835153</citedby><cites>FETCH-LOGICAL-c477t-6e82d5da697c38f62d141d3f1fd75a7df66de683bc5d7d34046df0742be835153</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><link.rule.ids>314,780,784,27924,27925</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/8063827$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><creatorcontrib>Crane, D I</creatorcontrib><creatorcontrib>Kalish, J E</creatorcontrib><creatorcontrib>Gould, S J</creatorcontrib><title>The Pichia pastoris PAS4 gene encodes a ubiquitin-conjugating enzyme required for peroxisome assembly</title><title>The Journal of biological chemistry</title><addtitle>J Biol Chem</addtitle><description>We report here the cloning and initial characterization of PAS4, a gene required for peroxisome assembly in the yeast Pichia
pastoris. The PAS4 gene encodes a 24-kDa protein (Pas4p) that is located on the cytoplasmic surface of peroxisomes and is
induced during peroxisome proliferation. Analysis of the Pas4p sequence revealed a high degree of similarity to ubiquitin-conjugating
enzymes, particularly in the region surrounding the putative active-site cysteine residue with which ubiquitin forms a thioester
bond. As expected for a ubiquitin-conjugating enzyme, substitution of alanine or serine for the conserved active-site cysteine
residue abolished PAS4 function. In addition, a small amount of a 32 kDa form of Pas4p (the predicted size of a Pas4p-ubiquitin
conjugate) was detected both in vivo and in vitro. This species was eliminated by reducing agents and was not detected in
the cysteine to alanine substitution mutant, suggesting that it is a Pas4p-ubiquitin conjugate. Using a yeast strain that
overexpresses a Myc-ubiquitin fusion protein, we demonstrate directly that this conjugate contains ubiquitin. We conclude
from these observations that PAS4 is a member of the ubiquitin-conjugating enzyme gene family and that one or more ubiquitination
reactions are required for peroxisome assembly.</description><subject>Amino Acid Sequence</subject><subject>Base Sequence</subject><subject>Cell Compartmentation</subject><subject>Enzyme Induction</subject><subject>Fungal Proteins</subject><subject>Genes, Fungal - genetics</subject><subject>Genes, myc - genetics</subject><subject>Genetic Complementation Test</subject><subject>Intracellular Membranes - chemistry</subject><subject>Ligases - analysis</subject><subject>Ligases - genetics</subject><subject>Ligases - metabolism</subject><subject>Microbodies - chemistry</subject><subject>Microbodies - physiology</subject><subject>Molecular Sequence Data</subject><subject>Mutation</subject><subject>Pichia - enzymology</subject><subject>Pichia - genetics</subject><subject>Pichia pastoris</subject><subject>Recombinant Fusion Proteins - metabolism</subject><subject>Sequence Homology, Amino Acid</subject><subject>Ubiquitin-Conjugating Enzymes</subject><subject>Ubiquitins - metabolism</subject><issn>0021-9258</issn><issn>1083-351X</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>1994</creationdate><recordtype>article</recordtype><sourceid>EIF</sourceid><recordid>eNqFUU1r3DAQFSUl2Sb9CQEdQmgPbjWWLMnHsPQLAg0khdyELI3XCmtrI61Jt7--2u6Sa-cyw7z3Zpg3hFwC-wQM5OfMWA1VWzf6A6iPHLRqK_6GLIBpXvEGHk_I4pVyRt7l_MRKiBZOyalmkutaLQg-DEjvghuCpRubtzGFTO9u7gVd4YQUJxc9Zmrp3IXnOWzDVLk4Pc0rW8pVwf_sRqQJC5bQ0z4musEUf4ccS9_mjGO33l2Qt71dZ3x_zOfk19cvD8vv1e3Pbz-WN7eVE0ptK4m69o23slWO617WHgR43kPvVWOV76X0KDXvXOOV54IJ6XumRN2hLhc3_JxcH-ZuUnyeMW_NGLLD9dpOGOdslJRSgGD_JYJsRcP0fmJzILoUc07Ym00Ko007A8zs_2Du9yabvckGlPn3B8OL7vK4YO5G9K-qo_EFvzrgQ1gNL8U704XoBhxNLYtemBrKSfwvsZSQUg</recordid><startdate>19940826</startdate><enddate>19940826</enddate><creator>Crane, D I</creator><creator>Kalish, J E</creator><creator>Gould, S J</creator><general>American Society for Biochemistry and Molecular Biology</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>7TM</scope><scope>M7N</scope><scope>7X8</scope></search><sort><creationdate>19940826</creationdate><title>The Pichia pastoris PAS4 gene encodes a ubiquitin-conjugating enzyme required for peroxisome assembly</title><author>Crane, D I ; Kalish, J E ; Gould, S J</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c477t-6e82d5da697c38f62d141d3f1fd75a7df66de683bc5d7d34046df0742be835153</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>1994</creationdate><topic>Amino Acid Sequence</topic><topic>Base Sequence</topic><topic>Cell Compartmentation</topic><topic>Enzyme Induction</topic><topic>Fungal Proteins</topic><topic>Genes, Fungal - genetics</topic><topic>Genes, myc - genetics</topic><topic>Genetic Complementation Test</topic><topic>Intracellular Membranes - chemistry</topic><topic>Ligases - analysis</topic><topic>Ligases - genetics</topic><topic>Ligases - metabolism</topic><topic>Microbodies - chemistry</topic><topic>Microbodies - physiology</topic><topic>Molecular Sequence Data</topic><topic>Mutation</topic><topic>Pichia - enzymology</topic><topic>Pichia - genetics</topic><topic>Pichia pastoris</topic><topic>Recombinant Fusion Proteins - metabolism</topic><topic>Sequence Homology, Amino Acid</topic><topic>Ubiquitin-Conjugating Enzymes</topic><topic>Ubiquitins - metabolism</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Crane, D I</creatorcontrib><creatorcontrib>Kalish, J E</creatorcontrib><creatorcontrib>Gould, S J</creatorcontrib><collection>Medline</collection><collection>MEDLINE</collection><collection>MEDLINE (Ovid)</collection><collection>MEDLINE</collection><collection>MEDLINE</collection><collection>PubMed</collection><collection>CrossRef</collection><collection>Nucleic Acids Abstracts</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>MEDLINE - Academic</collection><jtitle>The Journal of biological chemistry</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Crane, D I</au><au>Kalish, J E</au><au>Gould, S J</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>The Pichia pastoris PAS4 gene encodes a ubiquitin-conjugating enzyme required for peroxisome assembly</atitle><jtitle>The Journal of biological chemistry</jtitle><addtitle>J Biol Chem</addtitle><date>1994-08-26</date><risdate>1994</risdate><volume>269</volume><issue>34</issue><spage>21835</spage><epage>21844</epage><pages>21835-21844</pages><issn>0021-9258</issn><eissn>1083-351X</eissn><abstract>We report here the cloning and initial characterization of PAS4, a gene required for peroxisome assembly in the yeast Pichia
pastoris. The PAS4 gene encodes a 24-kDa protein (Pas4p) that is located on the cytoplasmic surface of peroxisomes and is
induced during peroxisome proliferation. Analysis of the Pas4p sequence revealed a high degree of similarity to ubiquitin-conjugating
enzymes, particularly in the region surrounding the putative active-site cysteine residue with which ubiquitin forms a thioester
bond. As expected for a ubiquitin-conjugating enzyme, substitution of alanine or serine for the conserved active-site cysteine
residue abolished PAS4 function. In addition, a small amount of a 32 kDa form of Pas4p (the predicted size of a Pas4p-ubiquitin
conjugate) was detected both in vivo and in vitro. This species was eliminated by reducing agents and was not detected in
the cysteine to alanine substitution mutant, suggesting that it is a Pas4p-ubiquitin conjugate. Using a yeast strain that
overexpresses a Myc-ubiquitin fusion protein, we demonstrate directly that this conjugate contains ubiquitin. We conclude
from these observations that PAS4 is a member of the ubiquitin-conjugating enzyme gene family and that one or more ubiquitination
reactions are required for peroxisome assembly.</abstract><cop>United States</cop><pub>American Society for Biochemistry and Molecular Biology</pub><pmid>8063827</pmid><doi>10.1016/s0021-9258(17)31879-3</doi><tpages>10</tpages><oa>free_for_read</oa></addata></record> |
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| ispartof | The Journal of biological chemistry, 1994-08, Vol.269 (34), p.21835-21844 |
| issn | 0021-9258 1083-351X |
| language | eng |
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| source | MEDLINE; Elektronische Zeitschriftenbibliothek - Frei zugängliche E-Journals; Alma/SFX Local Collection |
| subjects | Amino Acid Sequence Base Sequence Cell Compartmentation Enzyme Induction Fungal Proteins Genes, Fungal - genetics Genes, myc - genetics Genetic Complementation Test Intracellular Membranes - chemistry Ligases - analysis Ligases - genetics Ligases - metabolism Microbodies - chemistry Microbodies - physiology Molecular Sequence Data Mutation Pichia - enzymology Pichia - genetics Pichia pastoris Recombinant Fusion Proteins - metabolism Sequence Homology, Amino Acid Ubiquitin-Conjugating Enzymes Ubiquitins - metabolism |
| title | The Pichia pastoris PAS4 gene encodes a ubiquitin-conjugating enzyme required for peroxisome assembly |
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