The fukutin protein family – predicted enzymes modifying cell-surface molecules
1999; Elsevier BV; Volume: 9; Issue: 22 Linguagem: Inglês
10.1016/s0960-9822(00)80039-1
ISSN1879-0445
Autores Tópico(s)Endoplasmic Reticulum Stress and Disease
ResumoFukuyama type congenital muscular dystrophy (FCMD) is an autosomal recessive disorder that is observed predominantly in Japanese populations [[1]Arahata K Ishii H Hayashi YK Congenital muscular dystrophies.Curr Opin Neurol. 1995; 8: 385-390Crossref PubMed Scopus (26) Google Scholar]. Recently, the cause of this syndrome was discovered to be lesions in the gene encoding the protein fukutin; these lesions involve retroposon insertion and point mutations, which result in a truncated protein [[2]Kobayashi K Nakahori Y Miyake M Matsumura K Kondo-lida E Nomura Y et al.An ancient retrotransposal insertion causes Fukuyama-type congenital muscular dystrophy.Nature. 1998; 394: 388-392Crossref PubMed Scopus (0) Google Scholar]. Consistent with the phenotypic patterns of the disease, the fukutin mRNA has been found in skeletal muscles, heart, brain and pancreas [[2]Kobayashi K Nakahori Y Miyake M Matsumura K Kondo-lida E Nomura Y et al.An ancient retrotransposal insertion causes Fukuyama-type congenital muscular dystrophy.Nature. 1998; 394: 388-392Crossref PubMed Scopus (0) Google Scholar]. Fukutin contains a signal peptide and is localized to the Golgi and secretory granules [[2]Kobayashi K Nakahori Y Miyake M Matsumura K Kondo-lida E Nomura Y et al.An ancient retrotransposal insertion causes Fukuyama-type congenital muscular dystrophy.Nature. 1998; 394: 388-392Crossref PubMed Scopus (0) Google Scholar]. Here, we report on a detailed computer analysis of the fukutin protein sequence, resulting in the prediction that it is an enzyme that modifies cell-surface glycoproteins or glycolipids. A gapped BLASTP [[3]Altschul SF Madden TL Schaffer AA Zhang J Zhang Z Miller W Lipman DJ Gapped BLAST and PSI-BLAST: a new generation of protein database search programs.Nucleic Acids Res. 1997; 25: 3389Crossref PubMed Scopus (56937) Google Scholar] search of the non-redundant (NR) database at the National Center for Biotechnology Information using the fukutin sequence as the query revealed significant hits (with e-values < 10−9) not only to Caenorhabditis elegans proteins, such as T07D3.4 and T07A5.1, but also to the uncharacterized protein RP688 from the intracellular parasitic bacterium Rickettsia prowazekii (e < 10−4). The RP688 sequence was used for further analysis of this protein family by iteratively searching the NR database using the PSI-BLAST program, which was run with the cut-off of e = 0.001 [[3]Altschul SF Madden TL Schaffer AA Zhang J Zhang Z Miller W Lipman DJ Gapped BLAST and PSI-BLAST: a new generation of protein database search programs.Nucleic Acids Res. 1997; 25: 3389Crossref PubMed Scopus (56937) Google Scholar]. At convergence, not only fukutin and its C. elegans homologs, but also bacterial proteins involved in polysaccharide/phosphorylcholine modification and a yeast protein involved in mannosyl phosphorylation of oligosaccharides were retrieved from the database. Reverse searches with these sequences retrieved the original members of the fukutin family without any false positives. Fukutin, therefore, belongs to a family of proteins associated with the modification of the cell surface. A multiple alignment of the fukutin protein family was constructed using Gibbs sampling, as implemented in PROBE [[4]Neuwald AF Liu JS Lipman DJ Lawrence CE Extracting protein alignment models from the sequence database.Nucleic Acids Res. 1997; 25: 1665-1677Crossref PubMed Scopus (198) Google Scholar], in conjunction with the −m4 option of PSI-BLAST (Figure 1). The alignment shows prominent conservation in an amino-terminal block, followed by a weakly conserved carboxy-terminal region. The most notable feature of the amino-terminal region is the presence of the strictly conserved signature G[TS]hhGhhx4hhxaxxDxD (in single-letter amino acid code and in which ‘h’ is a hydrophobic amino acid, ‘a’ is an aromatic amino acid and ‘x’ denotes any amino acid). A pattern search with this motif recovers from the NR database the fukutin family in its entirety without any false positives. The carboxy-terminal region contains a motif with a conserved aspartate residue flanked by hydrophobic residues (Figure 1). Secondary structure prediction using the PHD program [[5]Rost B PHD: predicting one-dimensional protein structure by profile-based neural networks.Methods Enzymol. 1996; 266: 525-539Crossref PubMed Google Scholar] suggests a compact α/β fold for the fukutin domain (Figure 1). LicD2 from Streptococcus pneumoniae is involved in the addition of phosphorylcholine residues to lipoteichoic acid, an important component of the cell wall in Gram-positive bacteria [[6]Zhang JR Idanpaan-Heikkila I Fischer W Tuomanen EI Pneumococcal licD2 gene is involved in phosphorylcholine metabolism.Mol Microbiol. 1999; 31: 1477-1488Crossref PubMed Scopus (100) Google Scholar]. It has been proposed that LicD2 is an enzyme that catalyzes the transfer of phosphorylcholine to its teichoic acid substrates from a CDP-choline donor [[6]Zhang JR Idanpaan-Heikkila I Fischer W Tuomanen EI Pneumococcal licD2 gene is involved in phosphorylcholine metabolism.Mol Microbiol. 1999; 31: 1477-1488Crossref PubMed Scopus (100) Google Scholar]. Similarly, the Haemophilus influenzae homolog — LicD is required for the addition of phosphorylcholine to the Gram-negative bacterial lipopolysaccharide [[7]Weiser JN Shchepetov M Chong ST Decoration of lipopolysaccharide with phosphorylcholine: a phase-variable characteristic of Haemophilus influenzae.Infect Immun. 1997; 65: 943-950Crossref PubMed Google Scholar]. The yeast protein MNN4 participates in a similar reaction in which a mannosyl phosphate residue is added from a GDP–mannose donor to both N-linked and O-linked oligosaccharides on proteins ([[8]Jigami Y Odani T Mannosylphosphate transfer to yeast mannan.Biochim Biophys Acta. 1999; 1426: 335-345Crossref PubMed Scopus (152) Google Scholar] and references therein). These studies, taken together with the sequence conservation pattern, suggest that the fukutin family proteins are phosphoryl-ligand transferases. The presence of the DxD motif and a distal aspartate residue that is conserved in most of the sequences suggests that these enzymes coordinate a divalent cation, which is similar to a number of nucleotidyltransferases (for example, see [[9]Aravind L Leipe DD Koonin EV Toprim – a conserved catalytic domain in type IA and II topoisomerases, DnaG-type primases, OLD family nucleases and RecR proteins.Nucleic Acids Res. 1998; 26: 4205-4213Crossref PubMed Scopus (295) Google Scholar]). Drawing from knowledge of the functions of these proteins and the subcellular localization of fukutin, we predict that this protein modifies cell-surface molecules, most probably through the attachment of phosphoryl-sugar moieties. The brains of individuals with FCMD show an abnormal pattern of gangliosides [[10]Izumi T Hara K Ogawa T Osawa M Saito K Novo ML Fukuyama Y Takashima S Abnormality of cerebral gangliosides in Fukuyama type congenital muscular dystrophy.Brain Dev. 1995; 17: 33-37Abstract Full Text PDF PubMed Scopus (10) Google Scholar], which suggests that the transferase activity of fukutin could participate in one of the many steps of glycolipid modifications. Glycolipid modifications are an important factor in determining the adhesive properties of cells [[11]Lloyd KO Furukawa K Biosynthesis and functions of gangliosides: recent advances.Glycoconj J. 1998; 15: 627-636Crossref PubMed Scopus (113) Google Scholar]. Thus, fukutin might regulate neural migration and muscle organization by affecting the biogenesis of crucial adhesion molecules. In this regard, it might be of interest that disruption of the basement membrane, linked to an M-laminin deficiency, has been reported in FCMD patients [[12]Xu H Christmas P Wu XR Wewer UM Engvall E Defective muscle basement membrane and lack of M-laminin in the dystrophic dy/dy mouse.Proc Natl Acad Sci USA. 1994; 91: 5572-5576Crossref PubMed Scopus (236) Google Scholar]. All the bacterial homologs of fukutin are from pathogenic bacteria and experimental evidence from H. influenzae [[7]Weiser JN Shchepetov M Chong ST Decoration of lipopolysaccharide with phosphorylcholine: a phase-variable characteristic of Haemophilus influenzae.Infect Immun. 1997; 65: 943-950Crossref PubMed Google Scholar] and S. pneumoniae [[6]Zhang JR Idanpaan-Heikkila I Fischer W Tuomanen EI Pneumococcal licD2 gene is involved in phosphorylcholine metabolism.Mol Microbiol. 1999; 31: 1477-1488Crossref PubMed Scopus (100) Google Scholar] shows that the surface modifications catalyzed by these proteins (LicD and LicD2, respectively) are necessary for the adhesion of the bacteria to mammalian lung cells and pathogenesis. The sequence conservation described here, together with the apparent absence of fukutin family members in non-pathogenic bacteria, might suggest that the genes coding for these proteins have been horizontally transferred from eukaryotes to the pathogenic bacteria. A member of the fukutin family was detected in an expressed sequence tag from Trypanosoma brucei (AI077222), which suggests a possible role for this protein in the surface modifications of this eukaryotic pathogen (data not shown). The proposed function and the phylogenetic distribution of the fukutin family members resemble those of the glycosyl transferases of the Fringe family that are involved in the modification of surface molecules of the Notch pathway in development [[13]Yuan YP Schultz J Mlodzik M Bork P Secreted fringe-like signaling molecules may be glycosyltransferases.Cell. 1997; 88: 9-11Abstract Full Text Full Text PDF PubMed Scopus (101) Google Scholar]. Like the fukutin family proteins, Fringe homologs are seen in pathogenic but not in free-living bacteria. Thus horizontal gene transfer from eukaryotes might be a common theme in the origin of the enzymes involved in surface modification in bacterial pathogens, resulting in similarities in adhesion mechanisms between these bacteria and eukaryotic cells. L Aravind, National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland 20894 and Department of Biology, Texas A&M University, College Station, Texas 77843, USA. EV Koonin, National Center for Biotechnology Information, National Library of Medicine, National Institutes of Health, Bethesda, Maryland 20894, USA. E-mail: [email protected]
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