What Was the Set of Ubiquitin and UbiquitinLike C

Authors: Caroline Michelle Patrick Vourc’h Laurence Mignon Christian R Andres

Publish Date: 2009/05/19

Volume: 68, Issue: 6, Pages: 616-628

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Abstract

Ubiquitin Ubconjugating enzymes E2 are key enzymes in ubiquitination or Ublike modifications of proteins We searched for all proteins belonging to the E2 enzyme superfamily in seven species Homo sapiens Mus musculus Drosophila melanogaster Caenorhabditis elegans Schizosaccharomyces pombe Saccharomyces cerevisiae and Arabidopsis thaliana to identify families and to reconstruct each family’s phylogeny Our phylogenetic analysis of 207 genes led us to define 17 E2 families with 37 E2 genes in the human genome The subdivision of E2 into four classes did not correspond to the phylogenetic tree The sequence signature HPN histidine–proline–asparagine followed by a tryptophan residue at 16 up to 29 amino acids was highly conserved When present the active cysteine was found 7 to 8 amino acids from the Cterminal end of HPN The secondary structures were characterized by a canonical alpha/beta fold Only family 10 deviated from the common organization because the proteins were devoid of enzymatic activity Family 7 had an insertion between beta strands 1 and 2 families 3 5 and 14 had an insertion between the active cysteine and the conserved tryptophan The threedimensional data of these proteins highlight a strong structural conservation of the core domain Our analysis shows that the primitive eukaryote ancestor possessed a diversified set of E2 enzymes thus emphasizing the importance of the Ub pathway This comprehensive overview of E2 enzymes emphasizes the diversity and evolution of this superfamily and helps clarify the nomenclature and true orthologies A better understanding of the functions of these enzymes is necessary to decipher several human diseasesUbiquitination is characterized by a rapid and reversible posttranslational covalent fixation of ubiquitin Ub onto proteins Ciechanover 2006 This mechanism which is part of the specific proteindegradation pathway by the 26S proteasome plays an important role in targeting proteins as well as intracellular signalling It has an impact on many cellular functions such as DNA repair transcription signal transduction endocytosis and sorting Welchman et al 2005The ubiquitination of proteins requires three types of enzymes First Ubactivating enzymes E1 form a thioester bond with Ub in an adenosine triphosphate–dependent reaction Second Ubconjugating enzymes E2 or Ubc carry Ub and transfer it either directly to the substrate protein or to a third type of enzyme the Ub ligases E3 These latter enzymes facilitate the ligation of Ub to the target protein The possible intervention of E4 enzymes has been reported in the formation of polyubiquitin chains of lengths 4 Ub Koegl et al 1999 Although polyubiquitination generally signals a protein for degradation by the proteasome monoubiquitination is mainly involved in signaling itself Deubiquitination in contrast is performed by several deubiquitinating enzymes Wilkinson 1997Ub is a small protein of 76 amino acids and highly conserved in eukaryotes Ub binds to the target protein by way of an isopeptide link between the Cterminal glycine and a lysine residue of the protein and it appears that three conserved lysines within Ub are crucial for the formation of polyubiquitin chains Several proteins similar to Ub Ublike Ubl—such as SUMO123 NEDD8RUB1 ATG8 ATG12APG12 ISG15 FAU or URM1—can also be used in similar mechanisms Proteins tagged by SUMO small Ub modifier or NEDD8 are not recognized for degradation however they play a role in gene transcription activation protein localization and stabilization Each target–function combination has its own unique combination of E1 E2 and E3 enzymesThe human genome comprises eight genes encoding E1 Ub and Ubl activating enzymes Ensembl release 50 July 2008 and four genes encoding the two subunits of the E1 enzyme involved in sumoylation Supplementary Table 1 E3 enzymes are the most numerous with probably 1000 members Pickart 2004 They are the most specific in targeting the proteins for ubiquitination and can be divided into two main groups RING and HECT homologous to the E6AP carboxyl terminus proteins However the catalytic modules of these two families are unrelated in sequence or structure HECT domaincontaining E3 enzymes form intermediate thioesters with Ub at their active site cysteine before transferring Ub to substrates whereas most RING finger domaincontaining E3 enzymes act as scaffolds that bind to E2 enzymes and substrates simultaneously Özkan et al 2005Explanative structure showing in their 3D context the different residues mentioned in Fig 6 Loops are labeled L1 to L8 βstrands are labeled S1 to S4 and helices are labeled H1 to H4 with “h” for the generally conserved 3/10 helix Adapted from 3D structure of UBE2D2 PDB source E2SK and legended from Winn et al 2004According to the nature of the E2 enzyme the type of ubiquitination can be different as well For example in yeast only UBC13 type E2 enzymes are able to form polyubiquitin chains bound on Lys63 and the presence of a Ubconjugating enzyme variant UEV is necessary for this activity Andersen et al 2005

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