Biomedical subjects

T Korpela

Publications and source records attributed to T Korpela.

At least 37 records · Page 2Linked to original sources

PubMed1997

Molecular model of an alternative splice variant of human IL-4, IL-4 delta 2, a naturally occurring inhibitor of IL-4-stimulated T cell proliferation.

The molecular model of IL-4 delta 2, a naturally occurring splice variant of human IL-4 with exons 1, 3, and 4 in an open reading frame, is described. The second exon codes the main part of the long loop AB connected the helices A and B in parallel superposition. Therefore the hydrophobic core and the native fold of the rest part of IL-4 delta 2 molecule could be preserved without any significant changes only in the case of revolution of the helix A relative to other helices. In the result, the dominated a left-handed four-helix bundle structure of IL-4 with an up-up-down-down structural pattern is transformed to the IL-4 delta 2 structure with a down-up-down-down structural pattern.

Humans↗

PubMed1996

Expression and biochemical analyses of the recombinant potato virus X 25K movement protein.

The 25K movement protein (MP) of potato virus X (PVX) is encoded by the 5'-proximal gene of three overlapping MP genes forming a 'triple gene block'. The PVX 25K MP (putative NTPase-helicase) has been synthesized in Escherichia coli as a recombinant containing a six-histidine tag at the amino terminus. The His-tagged 25K protein was purified in a one-column Ni-chelate affinity chromatography procedure. In the absence of any other viral factors, this protein had obvious Mg2+-dependent ATPase activity, which was stimulated slightly (1.7-1.9-fold) by various polynucleotides. Like other viral proteins possessing ATPase-helicase motifs and many plant viral movement proteins, the PVX 25K MP was able to bind nucleic acids in vitro. The RNA binding activity of the 25K MP was pronounced only at very low salt concentrations and was independent of its ATPase activity.

Acid Anhydride Hydrolases↗

PubMed1996

Phosphoserine aminotransferase from Bacillus circulans subsp. alkalophilus: purification, gene cloning and sequencing.

Two peaks of aspartate aminotransferase (AspAT) catalytic activity were observed during DEAE chromatography of a protein extract from alkalophilic B. circulans. The enzyme purified from the major peak appeared to be not aspartate but phosphoserine aminotransferase (PSAT) with a considerably high AspAT side activity. The sequence of the enzyme N-terminus was determined, and the PSAT gene was cloned as two separate fragments. DNA sequencing revealed the open reading frame for the PSAT starting from TTG, putative ribosomal binding site and terminator of transcription. The PSAT gene encodes a protein of 361 amino acids (M(r) 39793) which shows moderate homology to other known phosphoserine aminotransferases (36-46% of identity, 60-64% of similarity). The PSAT from the alkalophile shares with all of them the consensus sequence pattern around the pyridoxal 5'-phosphate attachment site.

Amino Acid Sequence↗

PubMed1996

Structure of cytokine hydrophobic cores.

Diagrams were constructed that describe, using a common frame of reference, the interactions that occur within the hydrophobic cores of the left-handed four-helix bundles of interleukin-2, -4, -5, granulocyte/macrophage-, granulocyte-, macrophage-colony-stimulating factor, beta- and gamma-interferon, growth hormone, and leukemia inhibitory factor. Based on the assumption that the topologies of the ligand-receptor complexes are similar, analogous patterns were obtained from these diagrams and lead to the detailed comparison of the hydrophobic cores. These patterns were then used to obtain pairwise rigid-body superpositions among all of the four-helix bundles; values of the root-mean-square deviations (C alpha-atoms) over the four alpha-helices of the bundles range from 1.14 A to 3.22 A. Two distinct groups are formed after clustering based on structural superposition of the ten cytokines.

Cytokines↗

PubMed1996

Crystallization and preliminary X-ray analysis of phosphoserine aminotransferase from Bacillus circulans subsp. alkalophilus.

Recombinant phosphoserine aminotransferase (EC 2.6.1.52) from Bacillus circulans subsp. alkalophilus was crystallized at room temperature from 0.1 M sodium acetate buffer, pH 4.6, and 2% PEG 20000, using macroseeding techniques. The crystals diffract X-rays to at least 2.0 A nominal resolution. They belong to space group C2 with unit cell dimensions a = 93.2 A, b = 93.1 A, c = 45.6 A, alpha = 90.0 degrees, beta = 106.8 degrees, gamma = 90.0 degrees. A native data set to 2.3 A has been collected. Assuming an average packing density of the crystals, there is one monomer in the asymmetric unit, resulting in a calculated solvent content of 48.2%.

Bacillus↗

PubMed1996

Aspartate aminotransferase from an alkalophilic Bacillus contains an additional 20-amino acid extension at its functionally important N-terminus.

Aspartate aminotransferase (AspAT), responsible for a minor part of the total AspAT enzymic activity in alkalophilic Bacillus circulans, was purified, its N-terminal amino acid sequence was determined, and its gene was cloned as two separate fragments. DNA sequencing showed an open reading frame of 432 amino acids (M(r) 47,439) exhibiting moderately low homology with AspATs from other sources. Sequence alignment of the enzyme with chicken mitochondrial, chicken cytoplasmic and Escherichia coli AspATs was performed with the MULTALIN program and further optimized assuming that the three-dimensional structures of the proteins were conserved. The primary structure of the studied AspAT diverged markedly from the others in the catalytically important small domain and in a segment of 31 amino acids in the large domain. The functional N-terminal arm was about two times longer than those of AspATs from other sources. According to the molecular model, the unique regions of B. circulans AspAT are all located together, forming a continuous network of contacts. Additional contacts formed by the elongated N-terminal arm may result in some limitation of domain movements in the alkalophilic enzyme in comparison to in other known AspATs.

Amino Acid Sequence↗

PubMed1996

Conservative hydrophobic interdomain contacts of IFN-gamma remain in P17 matrix protein of HIV-1.

A constructed scheme of the surface layers containing helices C, D, and E' of various polypeptide chains which participate in the interdomain contacts in IFN-gamma demonstrated two sites of localization of the conservative hydrophobic amino acids. An analogous scheme of the interaction of helices B, C, and D in the p17 matrix protein of HIV-1 showed that the majority of the hydrophobic positions are similar. These data confirm the structural similarity between p17 and IFN-gamma.

Amino Acid Sequence↗

PubMed1995

Dimer structure as a minimum cooperative subunit of small heat-shock proteins.

Recently, it has been shown that small heat-shock proteins (Hsp25, Hsp27) are molecular chaperones. They bind to thermally unfolded proteins and can also assist refolding of denatured proteins. Mammalian small Hsps can form oligomeric structures of about 32 subunits. Until now, no data about cooperativity and stability of the interactions between the subunits of sHsps are available. To analyze these interactions we studied mouse Hsp25 and human Hsp27 by difference adiabatic scanning microcalorimetry (DASM) and circular dichroism (CD). Here we show that, according to DASM data, the minimum cooperatively melting structure is a sHsp-dimer. CD data indicate that Hsp25 major secondary structure, the beta-pleated conformation, is resistant to acidic influence up to pH 4.5 and, at neutral pH values, to heat treatment up to 60 degrees C. The melting pattern of Hsp25/27 bears resemblance to alpha-crystallins. CD data indicate similar secondary, tertiary and quaternary structures of the proteins compared. This finding is in agreement with the revealed homology of primary structure of these proteins and their common chaperone function.

Animals↗

PubMed1995

The role of histidine residues in the catalytic act of cyclomaltodextrin glucanotransferase from Bacillus circulans var. alkalophilus.

Our previous study on cyclomaltodextrin glucanotransferase (CGTase) by chemical modification implied the importance of one or two histidine residues in the cyclization reaction of the enzyme. Based on a computer modelled three-dimensional structure of the CGTase, five histidine residues were chosen as targets for the site-directed mutagenesis. The histidine residues 98, 140, 233 and 327 were replaced by aspartate and His-177 by proline using polymerase chain reaction-mediated techniques. The CGTase variants H98D, H140D, H233D and H327D resulted in a profound decrease in the cyclizing and amylolytic activities, while mutation H177P had little influence on the activities but affected the thermal stability and the width of the pH optimum. It is suggested that His-98 functions as (or as a significant part of) the subsite 2 for the binding of the substrate in CGTase and therefore H98D destabilizes the intermediate for cyclization, but does not markedly affect the hydrolytic reactions. Mutants H140D and H233D produced only minor amounts of alpha-cyclodextrin, did not exhibit substrate inhibition with maltotriose and showed non-Michaelis-Menten kinetics. It is proposed that the variants H140D, H233D and H327D cause steric hindrances near the active center, while mutation H177D has similar consequences on the same site spatially.

Bacillus↗

PubMed1995

Molecular modeling of the steric structure of the envelope F1 antigen of Yersinia pestis.

Steric structure of the envelope F1 protein of Yersinia pestis was reconstructed by computer modeling taking into account structural similarities between F1 and interleukins (IL)-1 alpha, -beta, -ra and by using the known atomic coordinates for huIL-1 beta obtained by the X-ray crystallography. Of 18 hydrophobic positions forming a hydrophobic core in all the proteins studied with the IL-1-like conformation, 15 positions are occupied by hydrophobic residues in F1 protein as well. Of 8 homologous positions occupied by the amino acid residues of similar charge in all huIL-1 alpha, -beta, -ra, 5 positions are conserved in F1 antigen. The B-cell epitope accessible to antibodies in polymeric F1 is exposed as an hydrophilic loop at the surface opposite to the C-terminal sequence, forming a conserved binding site with periplasmic molecular chaperones.

Amino Acid Sequence↗

PubMed1995

Modelling of steric structure of a periplasmic molecular chaperone Caf1M of Yersinia pestis, a prototype member of a subfamily with characteristic structural and functional features.

Steric structure of Caf1M, a periplasmic molecular chaperone of Yersinia pestis, was reconstructed by computer modelling based on a statistically significant primary structure homology between Caf1M and PapD protein from Escherichia coli, and using the known atomic coordinates obtained by the X-ray crystallography for PapD. In the three-dimensional model of Caf1M an accessory sequence between F1 and G1 beta-strands (as compared to PapD) can form a strain-specific part of the binding pocket of surface organell subunits. This accessory sequence decreases the depth of the binding pocket. The characteristic structural feature of the subfamily of periplasmic molecular chaperones with the accessory sequence (Caf1M subfamily) is the existence of exposed to a solvent Cys residues in F1 and G1 beta-strands which can form disulfide bond in the putative binding pocket. The characteristic functional feature of Caf1M subfamily is the chaperoning of more simple compositions of virulence-associated surface organells (in the case of Y. pestis a capsule consists of only F1 protein). Highly conserved R82 and D93, located at the domain surface remote from the putative subunit binding pocket, can participate in direct contacts with the conserved portion of molecular usher proteins.

Amino Acid Sequence↗

PubMed1995

Structural and functional homology between periplasmic bacterial molecular chaperones and small heat shock proteins.

The periplasmic Yersinia pestis molecular chaperone Caf1M belongs to a superfamily of bacterial proteins for one of which (PapD protein of Escherichia coli) the immunoglobulin-like fold was solved by X-ray analysis. The N-terminal domain of Caf1M was found to share a 20% amino acid sequence identity with an inclusion body-associated protein IbpB of Escherichia coli. One of the regions that was compared, was 32 amino acids long, and displayed more than 40% identity, probability of random coincidence was 1.2 x 10(-4). IbpB is involved in a superfamily of small heat shock proteins which fulfil the function of molecular chaperone. On the basis of the revealed homology, an immunoglobulin-like one-domain model of IbpB three-dimensional structure was designed which could be a prototype conformation of sHsp's. The structure suggested is in good agreement with the known experimental data obtained for different members of sHsp's superfamily.

Amino Acid Sequence↗

PubMed1995

Some new aspects of molecular mechanisms of cyclosporin A effect on immune response.

A few protein targets were found to display a specific high-affinity interaction with the immunosuppressant cyclosporin A (CsA): cytosolic cyclophilins (CyP)A, B, C, D, E containing from 122 to 174 amino acid residues in a polypeptide chain, and secreted forms of CyP; CyP-40, 40-kDa CsA-binding polypeptide complexed with steroid receptor (SR); CyP-related 150-kDa receptor of natural killer (NK) cells; interleukin 8 (IL-8); actin; a family of molecular chaperones hsp70 and P-glycoprotein (P-GP). All CyPs possess peptidyl-prolyl cis-trans isomerase activity (PPIase) and may serve as ATP-independent molecular chaperone proteins. The CsA-CyP complexes are specific inhibitors of Ca(2+)-and calmodulin-dependent protein phosphatase calcineurin (CaN). The inhibition of CaN blocks the activation of genes of IL-2, IL-2R, IL-4, etc. in T cells. In addition, immunosuppressive and/or antiinflammatory activity of CsA can be executed via CyP-40 and hsp 70 complexed with SR, and following the interaction with CyP-related receptor of NK and with IL-8. CsA binding to CyPC, P-GP and actin may throw light on the biochemical events leading to nephrotoxicity and graft vessel disease, two major side effects produced by CsA. The discovery of the interaction of human immunodeficiency virus type 1 (HIV-1) Gag protein with CyP and effective disruption of this interaction by CsA may be important for our understanding of the pathology caused by this immunosuppressive virus and will inspire therapeutic strategies to nip HIV in the bud. Bacterial immunophilins (ImPs) contribute to the virulence of pathogenic microorganisms. Elucidation of molecular mechanisms of microbial ImPs' action in the pathogenesis of bacterial infections may lead to new strategies for designing antibacterial drugs.

ATP Binding Cassette Transporter, Subfamily B, Mem↗

PubMed1994

Common structural patterns of cytokine outer surfaces.

Schemes of the four-helix bundle surfaces of interleukin-2, -4, -5, granulocyte/macrophage-, granulocyte-, macrophage-colony-stimulating factor, interferon-beta, -gamma and growth hormone were designed. All cytokines appeared to have the structurally similar "holes" on the surfaces. They were suggested to serve as a part of the main receptor-binding sites.

Cytokines↗

PubMed1994

Modelling of interhelical contacts in interferons-beta, -gamma, and dimeric interleukin-5.

On the basis of crystallographic data, the alpha-helices of interferon-beta as well as each domain of interferon-gamma are located in two layers. This results in a different contribution from the amino acids placing in the middle and border alpha-helices of the layer to the hydrophobic core of the molecule. A close analysis of the structure of dimeric interleukin-5 shows related arrangements of alpha-helices. The full schemes of the interhelix contacts of interferons-beta, -gamma, and dimeric interleukin-5 were constructed. Schemes for interleukin-5 show that extension of helices correlates with the dimer topology.

Computer Simulation↗

PubMed1994

Cyclophilin A and an antibody against cyclosporin A resemble each other in their binding sites.

The three-dimensional structures of cyclosporin A complexed with cyclophilin A or Fab fragment of a monoclonal antibody were compared. In these two complexes conformations of the fragment D-alanine8-N-methylvaline11 in cyclosporin A were in a good agreement. In addition, cyclophilin A and the Fab fragment had related arrangements of the aromatic amino acids in their binding sites, implying that antibody independently utilizes similar structural themes for binding cyclosporin A as cyclophilin A.

Amino Acid Isomerases↗

PubMed1993

Structural repeats in cyclosporin A.

The conformation of various dipeptides of free and cyclophilin A-bound cyclosporin A were compared. Cyclosporin A was shown to contain in both states conformationally duplicated fragments in the putative binding sites for cyclophilin A and phosphatase calcineurin.

Amino Acid Isomerases↗

PubMed1993

Comparison of conformations of cyclosporin A and macrolide FK506 fragments: localization of putative binding sites with phosphatase calcineurin.

The three-dimensional structures of two immunosuppressants, cyclosporin A and macrolide FK506, were compared. The sites N-methylglycine3-N-methylleucine4 and valine5-N-methylleucine6 of cyclosporin A were found to be similar to each other (the root-mean-square value was 0.29 A for six reference points of the main chain) and also to the site C17-C22 of FK506 (the root-mean-square values were 0.33 A and 0.13 A, respectively). We suggest these fragments of cyclosporin A and FK506 make a major contribution to the interaction of the immunosuppressants with the phosphatase calcineurin.

Amino Acid Sequence↗