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J Oda

Publications and source records attributed to J Oda.

At least 37 records · Page 2Linked to original sources
PubMed1998

Calcium ion-dependent reactivation of a Pseudomonas lipase by its specific modulating protein, LipB.

LipB, the lipase activator protein of Pseudomonas aeruginosa TE3285, was overproduced in Escherichia coli, and purified 4.9-fold over the crude extract in the presence of SDS. The purified LipB reactivated the lipase from P. aeruginosa TE3285 denatured with guanidine hydrochloride, and its reactivation did not involve multiple turnover. In this reactivation, a 1:1 complex between the lipase and LipB was detected in a cross-linking experiment, suggesting that LipB still binds to the lipase after the reactivation. Calcium ion was essential for the complex formation and the reactivation, and addition of EDTA caused inactivation of the reactivated lipase bound to LipB more rapidly than the native lipase. These findings suggest that LipB could affect the calcium binding to the lipase in the reactivation process. LipB was unable to reactivate lipases from other sources except Pseudomonas sp. 109; this lipase has an amino acid sequence which is 98% identical to that of the lipase from P. aeruginosa TE3285. Thus, it may be concluded that LipB specifically recognizes a unique structural element of the lipase.

Bacterial Proteins↗
PubMed1998

Random mutagenesis on the Pseudomonas lipase activator protein, LipB: exploring amino acid residues required for its function.

LipB, lipase activator protein from Pseudomonas aeruginosa TE3285, specifically recovers the enzymatic activity of denatured inactive lipase. To find important amino acid residues of LipB in this reactivation, random mutagenesis using error-prone PCR was performed on a gene encoding the functional region of LipB. The resultant DNA library was introduced into the lipase expression system using Escherichia coli, and LipB mutants lacking lipase activity were selected by two screening procedures. First, on agar plates containing tributyrin as a substrate for lipase, single colonies lacking active lipase secretion were selected as clones missing the active LipB. Second, to exclude nonsense and frameshift mutants, the molecular size of LipB in the given clones was confirmed by Western blotting. From the selected mutants, of which multiple residues are replaced, five single-residue substituted mutants were newly prepared. Consequently, Y99C, Y99H, S102R and R115C mutants formed no detectable complex with the lipase and lost the in vitro reactivation activity. In the case of Y99C and R115C, their single cysteine residue formed the intermolecular disulfide bridge. Thus, Tyr99 and Arg115 are likely to exist on the molecular surface of LipB, and are candidates for residues that make direct interaction with the denatured lipase in the reactivation process.

Bacterial Proteins↗
PubMed1998

ATP-dependent inactivation of Escherichia coli gamma-glutamylcysteine synthetase by L-glutamic acid gamma-monohydroxamate.

Incubation of Escherichia coli gamma-glutamylcysteine synthetase with L-glutamic acid gamma-monohydroxamate and ATP caused slow but irreversible inhibition of the enzyme, and more than 90% activity was lost in three days. The enzyme was not inactivated when ATP was absent or L-aspartic acid beta-monohydroxamate was substituted for L-glutamic acid gamma-monohydroxamate, suggesting that the inactivation process reflected a mechanism-based reaction of L-glutamic acid gamma-monohydroxamate and ATP.

Adenosine Triphosphate↗
PubMed1998

Molecular properties and activity of amino-terminal truncated forms of lipase activator protein.

Two mutant forms, which had truncated N-terminals, of lipase activator protein (LipB) from Pseudomonas aeruginosa TE3285 were prepared, and their molecular properties and activity were compared with those of the full-length form. A truncated LipB lacking its hydrophobic N-terminal 21 residues was dispersed homogeneously in solution, and could reactivate the stoichiometric amount of denatured lipase. In contrast, full-length LipB formed soluble aggregates, and reactivated less than an equimolar amount of the lipase even under the most suitable conditions. These findings suggest that some or all of the N-terminal 21 residues caused aggregation of the protein molecules, and prevented LipB from fully stoichiometric reactivation. A truncated LipB lacking the N-terminal 61 residues also reactivated denatured lipase, suggesting that the N-terminal 61-residue region of LipB is not involved in reactivation.

Amino Acid Sequence↗
PubMed1998

Aspirin renography in diagnosis of unilateral renovascular hypertension.

We previously proposed that aspirin can enhance the diagnostic sensitivity of renography with [123I] orthoiodohippurate (OIH) in patients with suspected unilateral renovascular hypertension (RVH). In this study we assessed the sensitivity and specificity of this method. Twenty-four patients, 14 with unilateral RVH and 10 with essential hypertension, were studied. For all patients with RVH, definitive diagnosis was based on the results of angiography and the response to renal arterial angioplasty after this study. For all patients with essential hypertension, we confirmed that there was little if any stenosis of the renal artery by digital subtraction angiography or Doppler sonography. Renography with [123I]OIH or 99mTc-mercaptoacetyltriglycine (MAG3) was done once before and once after the oral administration of aspirin (20 mg/kg). We considered renal blood flow to be decreased if the time to the peak in renography was 5 min or more, and defined the peak times of the kidneys as different if separated by 2 min or more. Unilateral RVH was diagnosed when both criteria were met. Renography before aspirin administration indicated RVH in 7 of the 14 patients with RVH, and renography after aspirin indicated RVH in 13 of the 14 patients. Of the 10 patients with essential hypertension, renography before and after aspirin administration showed no difference between the kidneys in 8 patients. Thus, aspirin renography increased the sensitivity from 50% to 93%, but did not change the specificity (80%) in the diagnosis of RVH. These results suggest that renography with [123I]OIH or 99mTc-MAG3 for the diagnosis of unilateral RVH is more sensitive when aspirin is used.

Adult↗
PubMed1998

Selection of severely head injured patients for mild hypothermia therapy.

OBJECT: The authors have analyzed the efficacy of inducing mild hypothermia (34 degrees C) in 62 severely head injured patients to control fulminant intracranial hypertension. METHODS: All 62 patients fulfilled the following criteria: 1)persistent intracranial pressure (ICP) greater than 20 mm Hg despite fluid restriction, hyperventilation, and high-dose barbiturate therapy; 2) an ICP lower than the mean arterial pressure; and 3) a Glasgow Coma Scale (GCS) score of 8 or less on admission. The patients were divided into three groups based on computerized tomography findings: extracerebral hematoma (34 patients with subdural and/or epidural hematoma), focal cerebral lesion (20 patients with localized brain contusion and/or intracerebral hematoma), and diffuse swelling (eight patients with no focal mass lesion). Mild hypothermia prevented ICP elevation in 35 (56.5%) of the 62 patients whose ICP was greater than 20 mm Hg despite conventional therapies. Among those 35 patients whose ICP was controlled by mild hypothermia, 12 (34.3%) achieved functional recovery (good outcome or moderate disability). However, functional recovery was observed in only five (10.9%) of the 46 patients whose ICP was greater than 40 mm Hg after conventional therapies. Of 40 patients with an admission GCS score of 5 to 8, there were 11 (27.5%) who achieved functional recovery. On the contrary, mild hypothermia was not effective in 22 patients with an admission GCS score of 3 or 4. In the patients with focal cerebral lesions, ICP was controlled by mild hypothermia in 17 patients (85%) and patient outcome was intimately related to the extent of the damage. Among 18 patients with extracerebral hematoma who had a midline shift of 9 to 12 mm, raised ICP could be successfully controlled by mild hypothermia in 16 patients (88.9%) and three (16.7%) achieved functional recovery. However, ICP could not be controlled in patients with extracerebral hematoma who had a midline shift of 13 mm or more. In patients with diffuse swelling, ICP elevation could not be prevented at all by mild hypothermia. CONCLUSIONS: The authors conclude that mild hypothermia is effective for preventing ICP elevation in patients without diffuse brain swelling in whom ICP remains higher than 20 mm Hg but less than 40 mm Hg after conventional therapies.

Adolescent↗
PubMed1997

Nicked multifunctional loop of glutathione synthetase still protects the catalytic intermediate.

A derivative of glutathione synthetase (GSHase) with the multifunctional loop cleaved (nicked GSHase) was compared to both a deletion mutant of the loop (loopless GSHase) and wild-type with the intact loop (wild-type GSHase). The loop had been shown to be in a closed state in order to protect a catalytic intermediate and accelerate the reaction. Data indicated that cleavage of the loop resulted in a drastic decrease in glutathione synthetic activity which was similar to the results for the loop deletion. Kinetic analyses indicated that the manipulations of the loop impaired the substrate affinity, especially for glycine, and also catalytic efficiency. The nicked loop did not accelerate the reaction as fast as the intact loop; however, the catalytic intermediate was protected from hydrolysis by the cleaved loop as effectively as by the intact loop. These results suggest that the fragmental loop assumed the closed state. High concentrations of ATP showed some inhibitory effects on wild-type GSHase, while both nicked and loopless GSHase were not inhibited, indicating that the fragments of the nicked loop functioned independently. In conclusion, it is postulated that the two fragments of the nicked loop independently assumed the closed state to protect the catalytic intermediate and have lost the ability to accelerate glutathione synthesis.

Adenosine Triphosphate↗
PubMed1997

Burn injuries in the 1995 Hanshin-Awaji earthquake.

In the 1995 Hanshin-Awaji earthquake, 504 deaths were listed as fire related, although many of the victims may have been crushed or suffocated before they were burned. Census data related to surviving burn victims, however, were unknown. This study was designed to examine the medical requirements of those burn patients following the earthquake. Medical records of 2718 patients with injuries admitted to 95 hospitals during the 15 days after the earthquake were retrospectively reviewed. Only 44 patients (1.9 per cent) were hospitalized with burns. Scalds with less than 20 per cent total burn surface area (TBSA) were mainly observed; flame burns from earthquake-associated fires were rare. Morbidity rates increased in patients over 40 years old. Associated injuries were observed in 11 cases. These included three soft tissue injuries, one rib, three spine, three pelvis and two extremity fractures, and two cases of crush syndrome. Intensive care was required for only 10 patients, five of whom were transferred to hospitals that were undamaged or outside the earthquake zone. No relationship was noted between the number of burned houses and that of hospitalized burn patients. These results suggest that the number of burn patients requiring medical care was less than might have been expected in view of the total number of fire-related deaths in this urban earthquake.

Burns↗
PubMed1997

Purification and characterization of tobacco pathogenesis-related protein PR-5d, an antifungal thaumatin-like protein.

Cultured tobacco cells accumulate several pathogenesis-related proteins. A neutral PR-5 protein, PR-5d, was purified to homogeneity from such cells. PR-5d has highly hydrophobic characteristics, but hydropathy analysis of its primary structure did not show a hydrophobic domain. In a series of bioassays, purified PR-5d showed inhibitory activity against several phytopathogenic and non-phytopathogenic fungi as do other members of the PR-5 protein family. To study the antifungal mechanism based on three dimensional structure of PR-5d, purified PR-5d was crystallized. The preliminary X-ray analysis of the crystal revealed that the crystals belong to space group C2, with cell dimensions a = 80.2 A, b = 63.8 A, c = 45.7 A, and beta = 107.2 degrees, and diffract at least 1.8 A resolution.

Amino Acid Sequence↗
PubMed1997

Analysis of 372 patients with Crush syndrome caused by the Hanshin-Awaji earthquake.

OBJECTIVE: To clarify clinical features and determine the severity of injuries in patients with crush syndrome in Hanshin-Awaji earthquake. METHODS: We retrospectively reviewed medical records of 6,107 patients hospitalized in 95 hospitals, and identified 372 patients with crush syndrome. RESULTS: The major sites of crush injury were in the lower extremities (74%), followed by the upper extremities (10%), and the trunk (9%). Pelvic fractures, limb fractures, and abdominal injuries were the most frequently associated injury. Patients with trunk compression and/or with abdominal injury had a higher mortality rate. A total of 50 patients (13.4%) died. The causes of death within 5 days after the earthquake were hypovolemia and hyperkalemia. Peak serum creatine kinase concentration increased with the number of crushed extremities. Mortality and the risk of acute renal failure were higher in patients with creatine kinase concentration more than 75,000 micro/L. CONCLUSIONS: Peak serum concentration of creatine kinase as well as the number of injured extremities serve to estimate the severity of crush syndrome.

Abdominal Injuries↗
PubMed1997

Analysis of 2,702 traumatized patients in the 1995 Hanshin-Awaji earthquake.

BACKGROUND: This study was undertaken to define the factors that affected the final outcome of trauma patients in the Hanshin-Awaji earthquake. METHODS: Medical records of patients admitted to 95 hospitals within or surrounding the affected area during the first 15 days after the quake were reviewed. RESULTS: There were 2,702 traumatized patients. One-third of the patients were transported to hospitals in the surrounding area and had a mortality rate of 3%. The remainder, who were treated in the affected hospitals, showed a significantly higher mortality rate (8%; p < 0.05). Intensive care was provided for 513 patients, most of whom suffered from crush syndrome or from injuries to vital organs; these patients had a high mortality rate. Patients with other types of injuries had a lower mortality rate. CONCLUSION: Crush syndrome and injuries to vital organs were potentially life-threatening. We believe that early transportation of such patients to undamaged hospitals with the ability to provide intensive care would have improved the survival rate.

Abbreviated Injury Scale↗
PubMed1996

A pseudo-michaelis quaternary complex in the reverse reaction of a ligase: structure of Escherichia coli B glutathione synthetase complexed with ADP, glutathione, and sulfate at 2.0 A resolution.

The crystal structure of glutathione synthetase from Escherichia coli B complexed with ADP, glutathione, and sulfate has been determined at 2.0 A resolution. Concerning the chemical similarity of sulfate and phosphate, this quaternary complex structure represents a pseudo enzyme-substrate complex in the reverse reaction and consequently allows us to understand the active site architecture of the E. coli glutathione synthetase. Two Mg2+ ions are coordinated with oxygen atoms from the alpha- and beta-phosphate groups of ADP and from the sulfate ion. The flexible loops, invisible in the unliganded or the binary and ternary complex structures, are fixed in the quaternary complex. The larger flexible loop (Ile226-Arg241) includes one turn of a 310-helix that comprises the binding site of the glycine moiety of GSH. The small loop (Gly164-Gly167) is involved in nucleotide binding and acts as a phosphate gripper. The side chains of Arg210 and Arg225 interact with the sulfate ion and the beta-phosphate moiety of ADP. Arg 210 is likely to interact with the carboxylate of the C-terminal gamma-glutamylcysteine in the substrate-binding form of the forward reaction. Other positively charged residues in the active site (Lys125 and Lys160) are involved in nucleotide binding, directing the phosphate groups to the right position for catalysis. Functional aspects of the active site architecture in the substrate-binding form are discussed.

Adenosine Diphosphate↗
PubMed1996

Crystallization and preliminary crystallographic study of asparagine synthetase from Escherichia coli.

Crystals of Escherichia coli asparagine synthetase have been obtained from 45% saturated ammonium sulfate solution at pH 7.5 by using its Cys-free (Cys51-->Ala, Cys315-->Ala) mutant. The crystals belong to space group P2(1), with cell dimensions a = 52.90(4), b = 126.2 (2), c = 52.8 (1) A and beta = 105.3(1) degrees. The self-rotation function specifies that there is one dimer in the asymmetric unit and the subunits are related by a non-crystallographic twofold axis. Complete data sets up to 2.7 A resolution have been collected on an R-AXIS IIc imaging-plate system.

Journal Article↗
PubMed1996

Gadolinium-enhanced MR imaging of the wrist in rheumatoid arthritis: value of fat suppression pulse sequences.

OBJECTIVE: To determine the usefulness of fat-suppressed gadolinium (Gd)-enhanced MR imaging of the wrist in patients with rheumatoid arthritis (RA). DESIGN AND PATIENTS: Fat-suppressed Gd-enhanced T1-weighted spin-echo (SE) images were obtained and compared with other standard techniques in 38 wrists of 27 patients (22-77 years) with RA. Scoring based on the degree of synovial enhancement of each joint was developed and the total scores (J-score) were correlated with radiographic stage, C-reactive protein (CRP), erythrocyte sedimentation rate (ESR), and symptomatic change in the follow-up study. RESULTS: Synovial proliferations showed marked enhancement in all the wrists. In addition, contrast enhancement in the bone marrow and tenosynovium was seen in 36 and eight wrists respectively. Fat-suppressed Gd-enhanced T1-weighted images demonstrated these abnormalities better than other techniques. The J-scores correlated well with values of CRP (P = 0.0034), but not with radiographic stages and ESR. CONCLUSION: Fat-suppressed Gd-enhanced T1-weighted SE images can clearly demonstrate most of the essential lesions in RA including the proliferative synovium, bone erosion, bone marrow inflammatory change, and tenosynovitis. Scoring based on the extent of Gd-enhancement of synovium can be useful in the assessment of the inflammatory status.

Adult↗
PubMed1996

Vascular occlusion and cartilage disorders in osteonecrosis of the femoral head in rats.

The aim of our study was to clarify the role of lateral epiphyseal vessels in the development of femoral head osteonecrosis in rats. We did this by performing histological and immunological studies on rats at the age that they are most prone to spontaneous osteonecrosis. We observed that obstruction of the lateral epiphyseal vessels occurred at the site of cartilage penetration. This obstruction preceded the development of osteonecrosis in adjacent areas of the femoral head. We propose that a similar mechanism may be responsible, in part, for the development of Perthes disease in man.

Animals↗
PubMed1996

Crystal structure of glutathione synthetase at optimal pH: domain architecture and structural similarity with other proteins.

The crystal structure of Escherichia coli B glutathione synthetase (GSHase) has been determined at the optimal catalytic condition pH 7.5. The most significant structural difference from the structure at pH 6.0 is the movement of the central domain towards the N-terminal domain almost as a rigid body. As a result of this movement, new interdomain and intersubunit polar interactions are formed which stabilize the dimeric structure further. The structure of GSHase at optimal pH was compared with 294 other known protein structures in terms of the spatial arrangements of secondary structural elements. Three enzymes (D-alanine: D-alanine ligase, succinyl-CoA synthetase and the biotin carboxylase subunit of acetyl-CoA carboxylase) were found to have structures similar to the ATP-binding site of GSHase, which extends across two domains. The ATP-binding sites in these four enzymes are composed of two antiparallel beta-sheets and are different from the classic mononucleotide-binding fold. Except for these proteins, no significant structural similarity was detected between GSHase and the other ATP-binding proteins. A structural motif in the N-terminal domain of GSHase has been found to be similar to the NAD-binding fold. This structural motif is shared by a number of other proteins that bind various negatively charged molecules.

Acetyl-CoA Carboxylase↗
PubMed1996

Parathyroid hormone-related protein (PTHrP) action in rat articular chondrocytes: comparison of PTH(1-34), PTHrP(1-34), PTHrP(1-141), PTHrP(100-114) and antisense oligonucleotides against PTHrP.

Parathyroid hormone-related protein (PTHrP) is thought to be an important autocrine/paracrine factor for chondrocyte metabolism since mice lacking the PTHrP gene exhibit abnormal cartilage development. To determine the biological role of PTHrP in chondrocytes, we first compared the agonist potency of human (h) PTHrP(1-34) with hPTH(1-34) in cultured rat articular chondrocytes. Neither hPTHrP(1-34) nor hPTH(1-34) altered basal DNA synthesis, but attenuated the stimulatory effect of transforming growth factor beta (TGF-beta). Both agents suppressed the expression of alpha(1) type II collagen mRNA in a dose-response fashion with the same potency. In addition, the action of exogenously added hPTHrP(1-34) and hPTH(1-34) on intracellular cAMP and [Ca2+]i levels was similar. We next compared the effect of PTHrP within its entire amino acid sequence (1-141). With regard to thymidine incorporation, alpha(1) type II collagen gene expression and accumulation of cAMP and [Ca2+]i level, there was no significant difference between hPTHrP(1-34) and hPTHrP(1-141). PTHrP C-terminal (100-114) did not show any function. To further investigate PTHrP function, intracellular PTHrP translation was inhibited by a transgene of antisense oligonucleotides against PTHrP. Antisense oligonucleotides decreased PTHrP mRNA translation, specifically inhibited DNA synthesis in control as well as TGF-beta-treated chondrocytes and enhanced alpha(1) type II collagen mRNA expression in TGF-beta-treated chondrocytes. These results suggest that there is no significant difference between exogenously added hPTH(1-34), hPTHrP(1-34) and PTHrP(1-141) with regard to the biological action of these agents, including cell growth, differentiation and second messenger pathway. However, the result of DNA synthesis in the antisense PTHrP-inhibition study suggests that intracellular PTHrP may have an as yet unknown biological role, in addition to a classical PTH/PTHrP receptor-mediated function in the rat articular chondrocyte.

Animals↗