Subdivision of the MDR superfamily of medium-chain dehydrogenases/reductases through iterative hidden Markov model refinement

<p>Abstract</p> <p>Background</p> <p>The Medium-chain Dehydrogenases/Reductases (MDR) form a protein superfamily whose size and complexity defeats traditional means of subclassification; it currently has over 15000 members in the databases, the pairwise sequence identity is typically around 25%, there are members from all kingdoms of life, the chain-lengths vary as does the oligomericity, and the members are partaking in a multitude of biological processes. There are profile hidden Markov models (HMMs) available for detecting MDR superfamily members, but none for determining which MDR family each protein belongs to. The current torrential influx of new sequence data enables elucidation of more and more protein families, and at an increasingly fine granularity. However, gathering good quality training data usually requires manual attention by experts and has therefore been the rate limiting step for expanding the number of available models.</p> <p>Results</p> <p>We have developed an automated algorithm for HMM refinement that produces stable and reliable models for protein families. This algorithm uses relationships found in data to generate confident seed sets. Using this algorithm we have produced HMMs for 86 distinct MDR families and 34 of their subfamilies which can be used in automated annotation of new sequences. We find that MDR forms with 2 Zn²⁺ions in general are dehydrogenases, while MDR forms with no Zn²⁺in general are reductases. Furthermore, in Bacteria MDRs without Zn²⁺are more frequent than those with Zn²⁺, while the opposite is true for eukaryotic MDRs, indicating that Zn²⁺has been recruited into the MDR superfamily after the initial life kingdom separations. We have also developed a web site <url>http://mdr-enzymes.org</url> that provides textual and numeric search against various characterised MDR family properties, as well as sequence scan functions for reliable classification of novel MDR sequences.</p> <p>Conclusions</p> <p>Our method of refinement can be readily applied to create stable and reliable HMMs for both MDR and other protein families, and to confidently subdivide large and complex protein superfamilies. HMMs created using this algorithm correspond to evolutionary entities, making resolution of overlapping models straightforward. The implementation and support scripts for running the algorithm on computer clusters are available as open source software, and the database files underlying the web site are freely downloadable. The web site also makes our findings directly useful also for non-bioinformaticians.</p

Human liver mitochondrial aldehyde dehydrogenase: a C-terminal segment positions and defines the structure corresponding to the one reported to differ in the Oriental enzyme variant

AbstractA C-terminal segment of mitochondrial human liver aldehyde dehydrogenase was characterized. The results prove that a central part of this segment largely but not completely agrees with a structure of a tryptic peptide previously reported for the same isoenzyme. This part corresponds to a segment that contains the exchanged residue in the functionally deficient Oriental variant of mitochondrial aldehyde dehydrogenase [(1984) Proc. Natl. Acad. Sci. USA 81, 258-261]. The data suggest important functions for the C-terminal region of aldehyde dehydrogenase, clarify previously inconsistent results, and establish this structure in the typical enzyme, including the position corresponding to the mutation in the functional variant.Mitochondrial isoenzymeAmino acid sequenceIsoenzyme differenceStructure-function relationshi

Human surfactant polypeptide SP-B Disulfide bridges, C-terminal end, and peptide analysis of the airway form

AbstractHuman hydrophobic surfactant polypeptide, SP-B, purified from lung tissue by exclusion chromatography in organic solvents, has been characterized. The polypeptide is 79 residues long, has a C-terminal methionine, and contains seven Cys residues. Native human SP-B lacks free thiol groups. Three intrachain disulfide bridges were defined, linking CysK to Cys77, Cys11 to Cys71 and Cys35 to Cys46. The remaining Cys48 is concluded to link the protein chains into homodimers via an interchain disulfide to its counterpart in a second SP-B polypeptide. These SS bridges are identical to those in the porcine form and confirm a consistant and unique disulfide pattern for SP-B polypeptides in general

Prokaryotic 20β-hydroxysteroid dehydrogenase is an enzyme of the ‘short-chain, non-metalloenzyme’ alcohol dehydrogenase type

AbstractThe primary structure of 20β-hydroxysteroid dehydrogenase from Streptomyces hydrogenans was determined after FPLC purification of a commercial preparation. Peptides obtained from different proteolytic cleavages were purified by reverse phase HPLC. The 255-residue structure deduced was found to be distantly homologous to those of Drosophila alcohol dehydrogenase and several other dehydrogenases, establishing that prokaryotic 20β-hydroxysteroid dehydrogenase as a member of the ‘short-chain alcohol dehydrogenase family’. With the enzymes characterized, the identity is greatest (31–34%) towards 4 other prokaryotic dehydrogenases, but the family also includes mammalian steroid and prostaglandin dehydrogenases. These enzymes are low in Cys and have a strictly conserved Tyr residue that appears to be important

Proteome analysis of vernix caseosa

To access publisher full text version of this article. Please click on the hyperlink in Additional Links fieldVernix caseosa (vernix) is a white creamy substance covering the skin of the fetus during the last trimester of pregnancy. The function of vernix has long been debated but no consensus has been reached. We here report a proteome analysis of vernix using two-dimensional gel electrophoresis, matrix-assisted laser desorption/ionization mass spectrometry and liquid chromatography coupled to tandem mass spectrometry. We have identified 41 proteins, of which 25 are novel to vernix. Notably, 39% of the identified vernix proteins are components of innate immunity, and 29% have direct antimicrobial properties. These results form a substantial contribution to the knowledge of vernix composition and demonstrate that antimicrobial protection of the fetus and the newborn child is a major and important function of vernix

Molecular and Cellular Effects of C-peptide—New Perspectives on an Old Peptide

New results present C-peptide as a biologically active peptide hormone in its own right. Although C-peptide is formed from proinsulin and cosecreted with insulin, it is a separate entity with biochemical and physiological characteristics that differ from those of insulin. There is direct evidence of stereospecific binding of C-peptide to a cell surface receptor, which is different from those for insulin and other related hormones. The C-peptide binding site is most likely a G–protein–coupled receptor. The association constant for C-peptide binding is approximately 3 × 109M-1. Saturation of the binding occurs already at a concentration of about 1 nM, which explains why C-peptide effects are not observed in healthy subjects. Binding of C-peptide results in activation of Ca2+ and MAPK-dependent pathways and stimulation of Na+,K+-ATPase and eNOS activities. The latter 2 enzymes are both deficient in several tissues in type 1 diabetes. There is some evidence that C-peptide, and insulin may interact synergistically on the insulin signaling pathway. Clinical evidence suggests that replacement of C-peptide, together with regular insulin therapy, may be beneficial in patients with type 1 diabetes and serve to retard or prevent the development of long-term complications

The p53 target protein Wig-1 binds hnRNP A2/B1 and RNA Helicase A via RNA

AbstractThe p53-induced Wig-1 gene encodes a double stranded RNA-binding zinc finger protein. We generated Saos-2 osteosarcoma cells expressing tetracycline-inducible Flag-tagged human Wig-1. Induction of Wig-1 expression by doxycycline inhibited cell growth in a long-term assay but did not cause any changes in cell cycle distribution nor increased fraction of apoptotic cells. Using co-immunoprecipitation and mass spectrometry, we identified two Wig-1-binding proteins, hnRNP A2/B1 and RNA Helicase A, both of which are involved in RNA processing. The binding was dependent on the presence of RNA. Our results establish a link between the p53 tumor suppressor and RNA processing via hnRNPA2/B1 and RNA Helicase A.Structured summaryMINT-6542926, MINT-6542899:WIG1 (uniprotkb:Q9HA38) physically interacts (MI:0218) with hnRNP A2/B1 (uniprotkb:P22626) by anti bait coimmunoprecipitation (MI:0006)MINT-6542945:RHA (uniprotkb:Q08211) physically interacts (MI:0218) with hnRNP A2/B1 (uniprotkb:P22626) by anti bait coimmunoprecipitation (MI:0006)MINT-6542918, MINT-6542891:WIG1 (uniprotkb:Q9HA38) physically interacts (MI:0218) with RHA (uniprotkb:Q08211) by anti bait coimmunoprecipitation (MI:0006)MINT-6542867:WIG1 (uniprotkb:Q9HA38) physically interacts (MI:0218) with RHA (uniprotkb:Q08211) by anti tag coimmunoprecipitation (MI:0007)MINT-6542879:WIG1 (uniprotkb:Q9HA38) physically interacts (MI:0218) with hnRNP A2/B1(uniprotkb:P22626) by anti tag coimmunoprecipitation (MI:0007