Biomedical subjects

Hiroshi Oyama

Publications and source records attributed to Hiroshi Oyama.

At least 19 recordsLinked to original sources

PubMed2006

A technique for identifying three diagnostic findings using association analysis.

In diagnosing diseases in clinical practice, a combination of three clinical findings is often used to represent each disease. This is largely because it is often difficult or impractical to assess for all possible combinations of symptoms and abnormal exam findings that occur in any particular disease. For most diseases, diagnostic triads are based on empirical observations. In this study, we determined diagnostic triads for chronic diseases using data mining procedures. We also verified the combinations' validity as well as our procedure for determining them. We used symptoms and examination findings from 477 patients with chronic diseases, collected as part of a 35-year longitudinal study begun in 1968. For each patient there were 295 items from examinations in internal medicine, dermatology, ophthalmology, dentistry and blood tests. We judged each item to be either normal or abnormal, and restricted the analysis to the abnormal findings. To analyze such an exhaustive assortment, we used the data mining technique of association analysis. The analysis generated three clinical findings for each disease. Diseases were defined based on blood tests. Searching through all 295 items to find the three most useful clinical findings would be impractical on a commodity PC. However, by excluding normal items, we were able to sufficiently reduce the total number of combinations so as to make combinatorial analysis on a PC feasible. In addition to more accurate diagnoses, we believe our technique can identify those diagnostic data that are more cost effective in terms of time and other resources required for their collection.

Data Collection↗

PubMed2006

Synergetic effects of pressure and chemical denaturant on protein unfolding: stability of a serine-type carboxyl protease, kumamolisin.

Kumamolisin, a serine carboxyl proteinase, is very stable and hardly denatured by single perturbation of a chemical denaturant (urea), pressure (<500 MPa) or temperature (<65 degrees C). In order to investigate the cooperative effects of these three denaturing agents, DSC, CD, intrinsic fluorescence, and fourth derivative UV absorbance were measured under various conditions. By application of pressure to kumamolisin in 8 M urea solution, substantial red-shift in the center of fluorescence emission spectral mass was observed, and the corresponding blue-shift was observed for two major peaks in fourth derivative UV absorbance, under the similar urea-containing conditions. The denaturation curves were analyzed on the basis of a simple two-state model in order to obtain thermodynamic parameters (DeltaV, DeltaG, and m values), and the combined effects of denaturing agents are discussed, with the special interest in the large cavity and neighboring Trp residue in kumamolisin.

Aspartic Acid Endopeptidases↗

PubMed2006

Physics-based simulation of surgical fields for preoperative strategic planning.

Although careful planning of surgical approach is a key for success of surgery, conventional planning and simulation tools cannot support detailed discussion. This issue is derived from the difficulty of estimating complex physical behavior of soft tissues provided by a series of surgical procedures like cutting and deformation. This paper proposes an adaptive physics-based framework that simulates both interactive cutting and accurate deformation on virtual bodies, and performs preoperative planning for supporting strategic discussion. We focus on limited use of the two models: A particle-based model and an FEM-based model considering required quality and performance in different situations. FEM-based deformation of incision accurately produces estimated surgical fields. Based on the framework, a strategic planning system was developed for supporting decision of surgical approach using 3D representation of the surgical fields. We applied clinical CT dataset of an aortic aneurysm case to the system. Some experiments and usability tests confirmed that the system contributes to grasping 3D shape and location of the target organs and performs detailed discussion on patient-specific surgical approaches.

Aortic Aneurysm↗

PubMed2006

Processing, catalytic activity and crystal structures of kumamolisin-As with an engineered active site.

Kumamolisin-As is an acid collagenase with a subtilisin-like fold. Its active site contains a unique catalytic triad, Ser278-Glu78-Asp82, and a putative transition-state stabilizing residue, Asp164. In this study, the mutants D164N and E78H/D164N were engineered in order to replace parts of the catalytic machinery of kumamolisin-As with the residues found in the equivalent positions in subtilisin. Unlike the wild-type and D164N proenzymes, which undergo instantaneous processing to produce their 37-kDa mature forms, the expressed E78H/D164N proenzyme exists as an equilibrated mixture of the nicked and intact forms of the precursor. X-ray crystallographic structures of the mature forms of the two mutants showed that, in each of them, the catalytic Ser278 makes direct hydrogen bonds with the side chain of Asn164. In addition, His78 of the double mutant is distant from Ser278 and Asp82, and the catalytic triad no longer exists. Consistent with these structural alterations around the active site, these mutants showed only low catalytic activity (relative k(cat) at pH 4.0 1.3% for D164N and 0.0001% for E78H/D164N). pH-dependent kinetic studies showed that the single D164N substitution did not significantly alter the logk(cat) vs. pH and log(k(cat)/Km) vs. pH profiles of the enzyme. In contrast, the double mutation resulted in a dramatic switch of the logk(cat) vs. pH profile to one that was consistent with catalysis by means of the Ser278-His78 dyad and Asn164, which may also account for the observed ligation/cleavage equilibrium of the precursor of E78H/D164N. These results corroborate the mechanistic importance of the glutamate-mediated catalytic triad and oxyanion-stabilizing aspartic acid residue for low-pH peptidase activity of the enzyme.

Aspartic Acid Endopeptidases↗

PubMed2005

Interaction model between elastic objects for haptic feedback considering collisions of soft tissue.

The simulation of organ-organ interaction is indispensable for practical and advanced medical VR simulator such as open surgery and indirect palpation. This paper describes a method to represent real-time interaction between elastic objects for accurate force feedback in medical VR simulation. The proposed model defines boundary deformation of colliding elements based on temporary surface forces calculated by temporary deformation. The model produces accurate deformation and force feedback considering collisions of objects as well as prevents unrealistic overlap of objects. A prototype simulator of rectal palpation is constructed on general desktop PC with a haptic device, PHANToM. The system allows users to feel different stiffness of a rear elastic object located behind another elastic object. The results of experiments confirmed the method expresses organ-organ interaction in real-time and produces realistic and perceivable force feedback.

Computer Simulation↗

PubMed2005

Catalytic residues and substrate specificity of scytalidoglutamic peptidase, the first member of the eqolisin in family (G1) of peptidases.

Scytalidoglutamic peptidase (SGP) is the first-discovered member of the eqolisin family of peptidases with a unique structure and a presumed novel catalytic dyad (E136 and Q53) [Fujinaga et al., PNAS 101 (2004) 3364-3369]. Mutants of SGP, E136A, Q53A, and Q53E lost both the autoprocessing and enzymatic activities of the wild-type enzyme. Coupled with the results from the structural analysis of SGP, Glu136 and Gln53 were identified as the catalytic residues. The substrate specificity of SGP is unique, particularly, in the preference at the P3 (basic amino acid), P1' (small a.a.), and P3' (basic a.a.) positions. Superior substrates and inhibitors have been synthesized for kinetic studies based on the results reported here. kcat, Km, and kcat/Km of SGP for D-Dap(MeNHBz)-GFKFF*ALRK(Dnp)-D-R-D-R were 34.8 s-1, 0.065 microM, and 535 microM-1 s-1, respectively. Ki of Ac-FKF-(3S,4S)-phenylstatinyl-LR-NH2 for SGP was 1.2x10(-10) M. Taken together, we can conclude that SGP has not only structural and catalytic novelties but also a unique subsite structure.

Catalysis↗

PubMed2005

Improving precise positioning of surgical robotic instruments by a three-side-view presentation system on telesurgery.

For faultless collaboration among the surgeon, surgical staffs, and surgical robots in telesurgery, communication must include environmental information of the remote operating room, such as behavior of robots and staffs, vital information of a patient, named supporting information, in addition to view of surgical field. "Surgical Cockpit System, " which is a telesurgery support system that has been developed by the authors, is mainly focused on supporting information exchange between remote sites. Live video presentation is important technology for Surgical Cockpit System. Visualization method to give precise location/posture of surgical instruments is indispensable for accurate control and faultless operation. In this paper, the authors propose three-side-view presentation method for precise location/posture control of surgical instruments in telesurgery. The experimental results show that the proposed method improved accurate positioning of a telemanipulator.

Humans↗

PubMed2005

Catalytic residues and substrate specificity of recombinant human tripeptidyl peptidase I (CLN2).

Tripeptidyl peptidase I (TTP-I), also known as CLN2, a member of the family of serine-carboxyl proteinases (S53), plays a crucial role in lysosomal protein degradation and a deficiency in this enzyme leads to fatal neurodegenerative disease. Recombinant human TPP-I and its mutants were analyzed in order to clarify the biochemical role of TPP-I and its mechanism of activity. Ser280, Glu77, and Asp81 were identified as the catalytic residues based on mutational analyses, inhibition studies, and sequence similarities with other family members. TPP-I hydrolyzed most effectively the peptide Ala-Arg-Phe*Nph-Arg-Leu (*, cleavage site) (k(cat)/K(m) = 2.94 microM(-1).s(-1)). The k(cat)/K(m) value for this substrate was 40 times higher than that for Ala-Ala-Phe-MCA. Coupled with other data, these results strongly suggest that the substrate-binding cleft of TPP-I is composed of only six subsites (S(3)-S(3)'). TPP-I prefers bulky and hydrophobic amino acid residues at the P(1) position and Ala, Arg, or Asp at the P(2) position. Hydrophilic interactions at the S(2) subsite are necessary for TPP-I, and this feature is unique among serine-carboxyl proteinases. TPP-I might have evolved from an ancestral gene in order to cleave, in cooperation with cathepsins, useless proteins in the lysosomal compartment.

Aminopeptidases↗

PubMed2005

Molecular cloning of the gene encoding Vibrio metalloproteinase vimelysin and isolation of a mutant with high stability in organic solvents.

Vimelysin is a unique metalloproteinase from Vibrio sp. T1800 exhibiting high activity at low temperature and high stability in organic solvents such as ethanol. A 1,821 bp open reading frame of the vimelysin gene encoded 607 amino acid residues consisting of an N-terminal pro-region, a mature enzyme, and a C-terminal pro-region. The mature enzyme region showed 80%, 57% and 35% sequence identity with the mature forms of vibriolysin from V. vulnificus, pseudolysin from Pseudomonas aeruginosa, and thermolysin from Bacillus thermoproteolyticus, respectively. The catalytic residues and zinc-binding motifs of metalloproteinases are well conserved in vimelysin. The vimelysin gene was expressed in E. coli JM109 cells and the recombinant enzyme was purified as a 38-kDa mature form from cell-free extracts. The purified recombinant enzyme is indistinguishable from the enzyme purified directly from Vibrio. To obtain mutants exhibiting higher stability in organic solvents, random mutations were introduced by error-prone PCR and 600 transformants were screened. The N123D mutant exhibits two times higher stability in organic solvents than the wild-type enzyme. A plausible mechanism for the stability of the N123D mutant in organic solvents was discussed based on homology models of vimelysin and the N123D mutant.

Bacterial Proteins↗

PubMed2005

MVL: medical VR simulation library.

In the last ten years, medical VR techniques have much progress and many simulators have been developed for education, planning, rehearsal and so on. On the other hand, developing a simulator takes much more labor and cost. In this paper, we propose MVL: Medical Virtual reality simulation Library, which supports simulation of several significant medical manipulations considering multiple organ interaction. The result of developing simulators using MVL confirmed validity about variety and developing cost.

Computer Simulation↗

PubMed2005

GenoCache: a Genomics and Cancer Chemoprevention portal.

To integrate cancer chemoprevention information on a genomics basis, we have built a Web site named GenoCache (Genomics and Cancer Chemoprevention) that would provide users with a molecular biological view of cancer chemoprevention. The site is organized in four viewpoints: substances, genes or gene products with their genetic variations, cells or tissues with various cancer development states, and "pathways" or sequential interactions between these objects. We linked the relevant genes to dbSNP.

Chemoprevention↗

PubMed2004

Crystallographic and biochemical investigations of kumamolisin-As, a serine-carboxyl peptidase with collagenase activity.

Kumamolisin-As (previously called ScpA) is the first known example of a collagenase from the sedolisin family (MEROPS S53). This enzyme is active at low pH and in elevated temperatures. In this study that used x-ray crystallographic and biochemical methods, we investigated the structural basis of the preference of this enzyme for collagen and the importance of a glutamate residue in the unique catalytic triad (Ser(278)-Glu(78)-Asp(82)) for enzymatic activity. Crystal structures of the uninhibited enzyme and its complex with a covalently bound inhibitor, N-acetyl-isoleucyl-prolyl-phenylalaninal, showed the occurrence of a narrow S2 pocket and a groove that encompasses the active site and is rich in negative charges. Limited endoproteolysis studies of bovine type-I collagen as well as kinetic studies using peptide libraries randomized at P1 and P1', showed very strong preference for arginine at the P1 position, which correlated very well with the presence of a negatively charged residue in the S1 pocket of the enzyme. All of these features, together with those predicted through comparisons with fiddler crab collagenase, a serine peptidase, rationalize the enzyme's preference for collagen. A comparison of the Arrhenius plots of the activities of kumamolisin-As with either collagen or peptides as substrates suggests that collagen should be relaxed before proteolysis can occur. The E78H mutant, in which the catalytic triad was engineered to resemble that of subtilisin, showed only 0.01% activity of the wild-type enzyme, and its structure revealed that Ser(278), His(78), and Asp(82) do not interact with each other; thus, the canonical catalytic triad is disrupted.

Amino Acid Sequence↗

PubMed2004

The molecular structure and catalytic mechanism of a novel carboxyl peptidase from Scytalidium lignicolum.

The molecular structure of the pepstatin-insensitive carboxyl peptidase from Scytalidium lignicolum, formerly known as scytalidopepsin B, was solved by multiple isomorphous replacement phasing methods and refined to an R factor of 0.230 (R(free) = 0.246) at 2.1-A resolution. In addition to the structure of the unbound peptidase, the structure of a product complex of cleaved angiotensin II bound in the active site of the enzyme was also determined. We propose the name scytalidocarboxyl peptidase B (SCP-B) for this enzyme. On the basis of conserved, catalytic residues identified at the active site, we suggest the name Eqolisin for the enzyme family. The previously uninvestigated SCP-B fold is that of a beta-sandwich; each sheet has seven antiparallel strands. A tripeptide product, Ala-Ile-His, bound in the active site of SCP-B has allowed for identification of the catalytic residues and the residues in subsites S1, S2, and S3, which are important for substrate binding. The most likely hydrolytic mechanism involves nucleophilic attack of a general base (Glu-136)-activated water (OH(-)) on the si-face of the scissile peptide carbonylcarbon atom to form a tetrahedral intermediate. Electrophilic assistance and oxyanion stabilization is provided by the side-chain amide of Gln-53. Protonation of the leaving-group nitrogen is accomplished by the general acid function of the protonated carboxyl group of Glu-136.

Amino Acid Sequence↗

PubMed2004

Two inhibitor molecules bound in the active site of Pseudomonas sedolisin: a model for the bi-product complex following cleavage of a peptide substrate.

High-resolution crystallographic analysis of a complex of the serine-carboxyl proteinase sedolisin with pseudo-iodotyrostatin revealed two molecules of this inhibitor bound in the active site of the enzyme, marking subsites from S3 to S3('). The mode of binding represents two products of the proteolytic reaction. Substrate specificity of sedolisin was investigated using peptide libraries and a new peptide substrate for sedolisin, MCA-Lys-Pro-Pro-Leu-Glu#Tyr-Arg-Leu-Gly-Lys(DNP)-Gly, was synthesized based on the results of the enzymatic and crystallographic studies and was shown to be efficiently cleaved by the enzyme. The kinetic parameters for the substrate, measured by the increase in fluorescence upon relief of quenching, were: k(cat)=73+/-5 s(-1), K(m)=0.12+/-0.011 microM, and k(cat)/K(m)=608+/-85 s(-1)microM(-1).

Binding Sites↗

PubMed2004

1.2 A crystal structure of the serine carboxyl proteinase pro-kumamolisin; structure of an intact pro-subtilase.

Kumamolisin, an extracellular proteinase derived from an acido/thermophilic Bacillus, belongs to the sedolisin family of endopeptidases characterized by a subtilisin-like fold and a Ser-Glu-Asp catalytic triad. In kumamolisin, the Asp82 carboxylate hydrogen bonds to Glu32-Trp129, which might act as a proton sink stabilizing the catalytic residues. The 1.2/1.3 A crystal structures of the Glu32-->Ala and Trp129-->Ala mutants show that both mutations affect the active-site conformation, causing a 95% activity decrease. In addition, the 1.2 A crystal structure of the Ser278-->Ala mutant of pro-kumamolisin was determined. The prodomain exhibits a half-beta sandwich core docking to the catalytic domain similarly as the equivalent subtilisin prodomains in their catalytic-domain complexes. This pro-kumamolisin structure displays, for the first time, the uncleaved linker segment running across the active site and connecting the prodomain with the properly folded catalytic domain. The structure strongly points to an initial intramolecular activation cleavage in subtilases, as presumed for pro-subtilisin and pro-furin.

Amino Acid Sequence↗

PubMed2004

Purification and characterization of alkaline serine proteinase from photosynthetic bacterium, Rubrivivax gelatinosus KDDS1.

In order to reduce the protein content of wastewater, photosynthetic bacteria producing proteinases were screened from wastewater of various sources and stocked in culture. An isolated strain, KDDS1, was identified as Rubrivivax gelatinosus, a purple nonsulfur bacterium that secretes proteinase under micro-aerobic conditions under light at 35 degrees C. Molecular weight of the purified enzyme was estimated to be 32.5 kDa. The enzyme showed the highest activity at 45 degrees C and pH 9.6, and the activity was completely inhibited by phenylmethyl sulfonyl fluoride (PMSF), but not by EDTA. The amino-terminal 24 amino acid sequence of the enzyme showed about 50% identity to those of serine proteinases from Pseudoalteromonas piscicida strain O-7 and Burkholderia pseudomallei. Thus, the enzyme from Rvi. gelatinosus KDDS1 was thought to be a serine-type proteinase. This was the first serine proteinase characterized from photosynthetic bacteria.

Amino Acid Sequence↗

PubMed2004

FEM-based interaction model between elastic objects for indirect palpation simulator.

Indirect palpation is required to examine lots of cases like breast cancer and prostate malignance. This paper proposes interaction model between elastic objects to simulate indirect palpation. The interaction is simulated by displacement of colliding elements based on normal stress derived from temporary displacement. The physics-based approach represents the difference of physical properties such as stiffness of colliding objects. Deformation and haptic reproduction is possible to be carried out in real time with two organ models consisting of roughly 200 nodal points. As an example, we developed a rectal palpation simulator based on the proposed method. The experiment using rectal palpation simulator confirmed that the method enables a user to perceive difference of stiffness of prostate model located behind rectum model indirectly.

Computer Simulation↗

PubMed2004

Physics-based preoperative approach planning using hybrid virtual bodies.

This paper proposes a hybrid model mixing geometry and volume data to improve representation of virtual bodies. This model applies object-oriented data models and rendering techniques to virtual organs, and enables both interactive VR simulation and detailed volume visualization of tissue of interest (e.g. coronary). Also, a physics-based framework interactively simulates estimated surgical fields which are used in preoperative discussion. Based on the proposed methods, a VR-based strategic planning system is developed. The system does not need high cost manual segmentation of patient dataset and efficiently supports planning of surgical approaches in cardiovascular surgery.

Humans↗