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T Ooi

Publications and source records attributed to T Ooi.

At least 55 records · Page 3Linked to original sources
PubMed1988

Pig heart calpastatin: identification of repetitive domain structures and anomalous behavior in polyacrylamide gel electrophoresis.

Isolation and nucleotide sequencing of the complementary DNA for pig heart calpastatin have been completed. The amino acid sequence of 713 residues predicted from the nucleotide sequence contains five domains, each composed of approximately 140 amino acid residues. A unique N-terminal domain is followed by four mutually homologous domains. The best fit alignment of these four domains gives residue identities between any two domains of 22.5-36.0%. The analysis of the sequence similarities by several methods also suggests the existence of additional shorter repeats at intervals of 60-80 residues. The calculated molecular weight of pig calpastatin of 713 amino acid residues (Mr 77,122) is significantly lower than the value of purified pig heart calpastatin (Mr 107,000) estimated by polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS-PAGE). The expression of the calpastatin genes in Escherichia coli and the detection of the translation products of 713, 366, and 140 amino acid residues by the specific anti-calpastatin antibody indicate that the products always migrate considerably slow on SDS-PAGE, giving an average of 1.53 for the ratio of the molecular weight estimated by SDS-PAGE to the value calculated from the amino acid sequences. It is most likely that the discrepancy in the molecular weight is caused by an anomalous behavior of calpastatin in SDS-PAGE.

Amino Acid Sequence↗
PubMed1988

Segmentation of a protein into structural elements: analysis and classification of segments.

A radial location measure (designated N14) derived from the X-ray crystallographic data represents an extent of exposure of an amino acid residue to solvent or location of the residue in a globular protein. A protein structure can be segmented along the chain by cutting at minima of a smoothed profile of N14. The segment so defined is a structural element traversing from a turn to the next turn and usually contains a regular secondary structure of alpha-helix or beta-strand in the middle. The characters of segments obtained for 87 proteins were examined in terms of four parameters: (1) the average angular moment (a quantity similar to the hydrophobic moment) at 40 degrees, M(40 degrees); (2) the average angular moment at 100 degrees, M(100 degrees); (3) the average value of location measures; and (4) the length of a segment. The segments were classified into 10 groups according to relative distances among the segments in the space spanned by the four parameters as components. Segments located in connecting regions (c segment) were expressed by three parameters: M(40 degrees), M(100 degrees), and the average value of location measures, and were classified into eight groups. Features of the segments were analyzed in terms of secondary structures, folding types of proteins, and occurrence of segment pairs. Protein structures are described by the arrangement of characterized segments and c segments. Cutting positions for the segmentation could be assigned from a smoothed profile calculated from its amino acid sequence, and the positions were in coincidence with the experimental ones with an accuracy of 77%. The significance of this approach to the description of protein structures in terms of segments is discussed.

Amino Acid Sequence↗
PubMed1988

Effects of hydrated water on protein unfolding.

The conformational stability of a protein in aqueous solution is described in terms of the thermodynamic properties such as unfolding Gibbs free energy, which is the difference in the free energy (Gibbs function) between the native and random conformations in solution. The properties are composed of two contributions, one from enthalpy due to intramolecular interactions among constituent atoms and chain entropy of the backbone and side chains, and the other from the hydrated water around a protein molecule. The hydration free energy and enthalpy at a given temperature for a protein of known three-dimensional structure can be calculated from the accessible surface areas of constituent atoms according to a method developed recently. Since the hydration free energy and enthalpy for random conformations are computed from those for an extended conformation, the thermodynamic properties of unfolding are evaluated quantitatively. The evaluated hydration properties for proteins of known transition temperature (Tm) and unfolding enthalpy (delta Hm) show an approximately linear dependence on the number of constituent heavy atoms. Since the unfolding free energy is zero at Tm, the enthalpy originating from interatomic interactions of a polypeptide chain and the chain entropy are evaluated from an experimental value of delta Hm and computed properties due to the hydrated water around the molecule at Tm. The chain enthalpy and entropy thus estimated are largely compensated by the hydration enthalpy and entropy, respectively, making the unfolding free energy and enthalpy relatively small. The computed temperature dependences of the unfolding free energy and enthalpy for RNase A, T4 lysozyme, and myoglobin showed a good agreement with the experimental ones.(ABSTRACT TRUNCATED AT 250 WORDS)

Kinetics↗
PubMed1988

Characteristic thermodynamic properties of hydrated water for 20 amino acid residues in globular proteins.

Thermodynamic properties associated with hydrated water of proteins of known three-dimensional structure were computed and average values of hydration free energy, enthalpy, and heat capacity of unfolding for every amino acid residue were obtained. Each amino acid residue had characteristic values; in particular, the quantities for a side chain reflected the character of the amino acid, while those for the main chain were more or less the same except for glycine, alanine, and proline. The major contribution to the quantities was from the end group(s) of a side chain. The following interesting features were found. 1) The hydration quantity of unfolding derived from the native and extended conformations for a protein was approximately equal to the sum of the corresponding average quantities of component amino acid residues in the protein. 2) The profile of a quantity such as hydration free energy of unfolding along the sequence computed from the accessible surface areas of the native and extended conformations showed a strong correlation with the profile obtained by allocating the average value for the amino acid residue at every position on the sequence. The correlation coefficients between two profiles for unfolding quantities of hydration, i.e., free energy, enthalpy, heat capacity, and free energy of side chain are 0.72, 0.62, 0.80, and 0.75, respectively. Thus, every amino acid residue in the native conformation of a globular protein seems to be located in such a position that a thermodynamic quantity for each residue is approximately equal to its average value.

Amino Acids↗
PubMed1988

Intermolecular interactions between protein and other molecules including hydration effects.

The structural aspects of protein functions, e.g., molecular recognition such as enzyme-substrate and antibody-antigen interactions, are elucidated in terms of dehydration and atomic interactions. When a protein interacts with some target molecule, water molecules at the interacting regions of both molecules are removed, with loss of the hydration free energy, but gaining atomic interactions between atoms of the contact sites in both molecules. The free energies of association originating from the dehydration and interactions between the atoms can be computed from changes in the accessible surface areas of the atoms involved. The free energy due to interactions between atomic groups at the contact sites is estimated as the sum of those estimated from the changes in the accessible surface area of 7 atomic groups, assuming that the interactions are proportional to the change of the area. The chain enthalpies and entropies evaluated from experimental thermodynamic properties and hydration quantities at the standard temperature for 10 proteins were available to determine the proportional constants for the atomic groups. This method was applied to the evaluation of association constants for the dimerization of proteins and the formation of proteolytic enzyme-inhibitor complexes, and the computed constants were in agreement with the experimental ones. However, the method is not accurate enough to account quantitatively for the change in the thermal stability of mutants of T4 lysozyme. Nevertheless, this method provides a way to elucidate the interactions between molecules in solution.

Proteins↗
PubMed1988

Chain reversals in model peptides: studies of cystine-containing cyclic peptides. II. Effects of valyl residues and possible i-to-(i + 3) attractive ionic interactions on cyclization of [Cys1], [Cys6] hexapeptides.

The synthesis of four N-acetyl N'-methylamide cystine-containing hexapeptides, CVPGVC, CGVVGC, CKPGEC, and CEPGKC, is described. These were used in disulfide-exchange reactions with the peptide CVPGGC as the formal oxidant. The relative propensities for peptide cyclization were thus deduced, and the tendency toward the formation of chain-reversal conformations was established quantitatively. An additional peptide, CVVVVC, was prepared but was never obtained as the cyclic monomer, demonstrating that the formation of chain-reversals in this peptide was of very low probability. Incorporation of pairs of valyl residues decreased the ease of cyclization, but it appeared that conformational flexibility in the cystine-containing hexapeptides may have compensated for substitutions which would have been expected to hinder the adoption of certain beta-turn conformations. The peptides containing ionic residues were cyclized more readily than expected, and this process was relatively insensitive to salt concentration. This observation is discussed with regard to the stabilization of beta-turns by i-to-(i + 3) ionic interactions in peptides and proteins. A method for blocking thiols was introduced as an improvement in the analysis of the equilibrium mixtures.

Amino Acid Sequence↗
PubMed1988

[Combination intra-arterial chemotherapy with degradable starch microspheres and mitomycin C against inoperable hepatic metastases].

Intra-arterial hepatic infusion chemotherapy combined with degradable starch microspheres (DSM) and mitomycin C (MMC) was performed for 9 patients with inoperable hepatic metastases from alimentary tract primary cancer. DSM, 45 +/- 5 micron in average diameter, produces temporary obstruction of arterial blood flow in the arterio-capillary bed and are subsequently degraded by serum amylase with T 1/2 of about 30 min. This intra-arterial treatment was repeated 2.3 times on the average. The average dose of DSM in a single infusion was 721 +/- 194 mg and the average total dose of MMC was 34.4 +/- 22.3 mg. Antitumor effects were evaluated in terms of tumor regression measured by CT scan and sonography. An objective tumor response was shown in 4/9 patients (44.4%): PR, 2/9; MR, 2/9. Elevated serum CEA levels of more than 3.0ng/ml were decreased in 7/8 patients (87.5%). Marked declines in the CEA level of more than 50% were observed in 4/8 patients (50%). Nausea and vomiting as well as abdominal pain were experienced in 8/21 treatments (38.1%) and 5/21 (23.8%), respectively. Furthermore, fever (2/21 : 9.5%), hepatic dysfunction (2/21 : 9.5%), and leukopenia (1/21 : 4.8%) were observed. All these side effects, however, were mild and transient. Thus, these results suggest that combined intra-arterial administration of DSM and MMC favorably enhances the antitumor efficacy of MMC.

Carcinoma, Hepatocellular↗
PubMed1988

[Repeated intra-arterial chemotherapy combined with mitomycin C and degradable starch microspheres in inoperable metastatic hepatic cancer].

Intra-arterial chemotherapy combined with mitomycin C (MMC) and degradable starch microspheres (DSM) was prescribed repeatedly for 14 patients with inoperable hepatic metastasis. This intra-arterial treatment was repeated 2.7 times on the average. The average dose of DSM and MMC in a single infusion was 685 +/- 201 mg and 13.8 +/- 3.8 mg, respectively. To analyse the degradation time of DSM, MMC concentrations in the peripheral blood were measured by HPLC method. RI-angiography using 99mTc macroaggregated albumin(MAA) was performed to estimate the hemodynamic changes in the liver. Antitumor efficacy was evaluated in terms of tumor regression measured by CT scan. An objective tumor response was observed in 9/14 patients(64.2%): CR 1/14; PR 6/14; MR 2/14. Elevated CEA levels were decreased in 11/12 patients (91.7%). Based on the peripheral MMC blood levels after combined infusion with DSM and MMC, an occlusion of intrahepatic vessels with DSM continued at least for 30 minutes. Again, RI-angiography with 99mTc-MAA plus DSM revealed the increased accumulation in the tumor, compared to 99mTc-MAA only. Side effects possibly attributable to DSM were observed in 14/38 treatments, though they were slight and temporary. Thus, these results indicate that this combined intra-arterial infusion of DSM and MMC achieve, higher regional selectivity.

Combined Modality Therapy↗
PubMed1987

Homology between mammalian DNA polymerase beta and terminal deoxynucleotidyltransferase.

Nucleotide sequence analysis of the cDNA and the genomic clones for rat DNA polymerase beta revealed the existence of a 1,005-base pair open reading frame capable of encoding a Mr = 38,269 polypeptide of 335 amino acid residues. The region of 174 amino acid residues between the 42nd and 215th residues of the DNA polymerase beta polypeptide has extensive amino acid sequence homology with the region between the 195th and 366th residues of human terminal deoxynucleotidyltransferase. The two enzymes share extensive homology not only in primary structures but also in the computer-derived higher structures in these particular regions. The genes for DNA polymerase beta and terminal deoxynucleotidyltransferase are proposed to be derived from a common ancestral DNA polymerase gene.

Amino Acid Sequence↗
PubMed1987

Examination of protein sequence homologies: IV. Twenty-seven bacterial ferredoxins.

Sequence homologies of 27 bacterial ferredoxins were examined using a computer program that quantitatively evaluates extent of similarity as a correlation coefficient. The results of a similarity search among the sequences demonstrated that the basal sequence consists of a pair of extremely similar segments of 26 amino acids connected by a three-amino acid group. The segment pairs, which would have arisen from gene duplication, are termed the first and second units. Because of the gene duplication, the connector sequence appears to have been introduced as a structurally important chain reversal. Each of the two units contains four cysteine residues, which are inserted one by one among seven, two, two, three, and eight amino acid alignments, respectively. The bacterial ferredoxins were categorized with regard to basal constitution as follows: group 1, in which both units closely conform to the basal structure; group 2, in which the second unit is modified in a characteristic manner among members; group 3, in which the first unit is modified in a characteristic manner, while the conforming second unit is accompanied by a long accessory sequence; group 4, in which there are modifications before and/or after the units, of which the respective central domains remain nearly intact; and group 5, where only the former of two Fe:S cluster ligation sets of four cysteines is estimated to remain intact, whereas the latter set is extremely modified. It is noteworthy that throughout all bacterial ferredoxins, one of two cysteine sets never fails to be completely intact and, moreover, the connector of three amino acids also exists intact. Based on this grouping and on the correspondences among the groups, average correlation coefficients among all members were computed, and the respective evolutionary relationships were examined. The results supported the proposition that transposition had occurred in the Azotobacter-type ferredoxins of group 3.

Amino Acid Sequence↗
PubMed1987

Accessible surface areas as a measure of the thermodynamic parameters of hydration of peptides.

A method is described for the inclusion of the effects of hydration in empirical conformational energy computations on polypeptides. The free energy of hydration is composed of additive contributions of various functional groups. The hydration of each group is assumed to be proportional to the accessible surface area of the group. The constants of proportionality, representing the free energy of hydration per unit area of accessible surface, have been evaluated for seven classes of groups (occurring in peptides) by least-squares fitting to experimental free energies of solution of small monofunctional aliphatic and aromatic molecules. The same method has also been applied to the modeling of the enthalpy and heat capacity of hydration, each of which is computed from the accessible surface area.

Amino Acids↗
PubMed1987

Detection of weak sequence homology of proteins for tertiary structure prediction.

Multiple measures of similarity were employed to detect weak homologies among protein sequences (e.g., below 30% residue identity). A set of thresholds was empirically determined, by using sample proteins of known structure, so as to select only correct pairs of sequences; correct or incorrect alignment of sequences was judged by direct comparison of corresponding conformations. The empirical criterion thus set up is applicable to the prediction of a protein structure when the structure of the other protein in the pair is known. We searched all the combinations between 84 proteins of known structure and 4610 proteins stored in a sequence database, and found about 4000 pairs of sequences which satisfied the criterion. However, after excluding such pairs of proteins that belong to the same family or superfamily, the number of pairs remaining was reduced to only 19. The reliability of these data for structural prediction is discussed.

Amino Acid Sequence↗
PubMed1986

Amino acid sequence homology applied to the prediction of protein secondary structures, and joint prediction with existing methods.

The assumption that homologous segments in different proteins may share a similar conformation is applied to the prediction of secondary structures in proteins. Sequences homologous to a target protein are searched, without allowing any gap, and compared against a number of reference proteins of known three-dimensional structure, and then a conformational state (alpha, beta or coil) for each residue of the protein is predicted by looking at the secondary structure of corresponding homologous segments. This prediction is done in a statistical rather than 'deterministic' way, by assigning the most probable conformation state among homologous data to each residue site of a target protein. A test application for 22 sample proteins yields 60% correctness on the average, a better value in comparison with two other existing methods. Joint prediction combining three methods into one is shown to increase the reliability up to 70%, when only the regions identically predicted with the three methods are taken into account. Application of the present method to 10 proteins of unknown structure is demonstrated.

Amino Acid Sequence↗
PubMed1986

Examination of protein sequence homologies: III. Ribosomal protein YS25 from Saccharomyces cerevisiae and its counterparts from Schizosaccharomyces pombe, rat liver, and Escherichia coli.

The sequences of the ribosomal proteins YS25, SP-S28, RL-S21, and Ec-S6, from Saccharomyces cerevisiae, Schizosaccharomyces pombe, rat liver, and Escherichia coli, respectively, have been examined using a computer program that searches for homologous tertiary structures. Matrices of comparisons among the eukaryotic sequences show that they match each other sequentially without any internal gaps. The average values of the correlation coefficients obtained from the comparison matrices are higher for the first halves of the sequences than for the latter halves. This result suggests that the first halves of the sequences may represent a more important domain than the latter halves. The comparison matrices between the eukaryotic and bacterial sequences of ribosomal proteins, however, do not show sequentially arranged homology, though there are six well-matching segments arranged in different orders in the two types of sequences. This implies that the eukaryotic sequences of the ribosomal protein were reconstituted by two internal transpositions and six deletions of 4-12 residues each from the ancestral sequence during the divergence between bacterial and eukaryotic genes. These findings may give insight into structural and quantitative studies of evolutionary divergence between eukaryotes and prokaryotes.

Amino Acid Sequence↗
PubMed1986

Radial locations of amino acid residues in a globular protein: correlation with the sequence.

The location measure of a residue in a globular protein is defined as the number of C alpha atoms surrounding the residue located within a sphere of the radius of 14 A. This quantity is a measure of the exposure of a residue to solvent, and is related closely to the distance from the center of mass of a protein. In this work, the experimental value for each residue of a protein is obtained from the X-ray crystallographic data, and the quantity is also calculated from the amino acid sequence data by applying an empirical parameter set to it. The correlation between the experimental and computed quantities is as high as 0.50 on the average over 92 proteins of known three-dimensional structure. Therefore, the location measure of every residue in a globular protein is predictable with good accuracy from the sequence.

Amino Acid Sequence↗
PubMed1986

The folding type of a protein is relevant to the amino acid composition.

The folding types of 135 proteins, the three-dimensional structures of which are known, were analyzed in terms of the amino acid composition. The amino acid composition of a protein was expressed as a point in a multidimensional space spanned with 20 axes, on which the corresponding contents of 20 amino acids in the protein were represented. The distribution pattern of proteins in this composition space was examined in relation to five folding types, alpha, beta, alpha/beta, alpha + beta, and irregular type. The results show that amino acid compositions of the alpha, beta, and alpha/beta types are located in different regions in the composition space, thus allowing distinct separation of proteins depending on the folding types. The points representing proteins of the alpha + beta and irregular types, however, are widely scattered in the space, and the existing regions overlap with those of the other folding types. A simple method of utilizing the "distance" in the space was found to be convenient for classification of proteins into the five folding types. The assignment of the folding type with this method gave an accuracy of 70% in the coincidence with the experimental data.

Amino Acids↗