Biomedical subjects

Atsushi Kurata

Publications and source records attributed to Atsushi Kurata.

6 recordsLinked to original sources

PubMed2007

Extremely high alkaline protease from a deep-subsurface bacterium, Alkaliphilus transvaalensis.

A new high-alkaline protease (ALTP) was purified to homogeneity from a culture of the strictly anaerobic and extremely alkaliphilic Alkaliphilus transvaalensis. The molecular mass was 30 kDa on sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The enzyme showed the maximal caseinolytic activity higher than pH 12.6 in KCl-NaOH buffer at 40 degrees C. Hydrolysis of the oxidized insulin B-chain followed by mass spectrometric analysis of the cleaved products revealed that as many as 24 of the total 29 peptide bonds are hydrolyzed in a block-cutting manner, suggesting that ALTP has a widespread proteolytic functions. Calcium ion had no effect on the activity and stability of ALTP, unlike known subtilisins. The deduced amino acid sequence of the enzyme comprised 279 amino acids plus 97 prepropeptide amino acids. The amino acid sequence of mature ALTP was confirmed by capillary liquid chromatography coupled to tandem mass spectrometry, which was the 93% coverage of the deduced amino acid sequence. The mature enzyme showed moderate homology to subtilisin LD1 from the alkaliphilic Bacillus sp. strain KSM-LD1 with 64% identity, and both enzymes formed a new subcluster at an intermediate position among true subtilisins and high-alkaline proteases in a phylogenetic tree of subtilase family A. ALTP is the first high-alkaline protease reported from a strict anaerobe in this family.

Amino Acid Sequence↗

PubMed2005

2-Haloacrylate reductase, a novel enzyme of the medium chain dehydrogenase/reductase superfamily that catalyzes the reduction of a carbon-carbon double bond of unsaturated organohalogen compounds.

A soil bacterium, Burkholderia sp. WS, grows on 2-chloroacrylate as the sole carbon source. To identify the enzymes metabolizing 2-chloroacrylate, we carried out comparative two-dimensional gel electrophoresis of the proteins from 2-chloroacrylate- and lactate-grown bacterial cells. As a result, we found that a protein named CAA43 was inducibly synthesized when the cells were grown on 2-chloroacrylate. The CAA43 gene was cloned and shown to encode a protein of 333 amino acid residues (M(r) 35,788) that shared a significant sequence similarity with NADPH-dependent quinone oxidoreductase from Escherichia coli (38.2% identity). CAA43 was overproduced in E. coli and purified to homogeneity. The purified protein catalyzed the NADPH-dependent reduction of the carbon-carbon double bond of 2-chloroacrylate to produce (S)-2-chloropropionate, which is probably further metabolized to (R)-lactate by (S)-2-haloacid dehalogenase in Burkholderia sp. WS. NADH did not serve as a reductant. Despite the sequence similarity to quinone oxidoreductases, CAA43 did not act on 1,4-benzoquinone and 1,4-naphthoquinone. 2-Chloroacrylate analogs, such as acrylate and methacrylate, were also inert as the substrates. In contrast, 2-bromoacrylate served as the substrate. Thus, we named this novel enzyme 2-haloacrylate reductase. This study revealed a new pathway for the degradation of unsaturated organohalogen compounds. It is also notable that the enzyme is useful for the production of (S)-2-chloropropionate, which is used for the industrial production of aryloxyphenoxypropionic acid herbicides.

Acrylates↗

PubMed2005

Inflammatory cells in the formation of tumor-related sarcoid reactions.

Tumor-related sarcoid reactions were analyzed in 14 lymph nodes in comparison with sarcoidosis using immunohistochemical markers to lymphocytes (CD3, CD4, CD8, and CD20), myeloid-related protein (MRP) 8 and MRP14 (S100A8 and S100A9), angiotensin I-converting enzyme (CD143), and mature or immature dendritic cells (S100, HLA-DR, fascin, CD83, and CD1a). We found that solitary epithelioid cell granuloma (ECG) first occur between lymph sinus and T-zone and that multiple ECGs mainly occur within T-zone, whereas confluent types often occupy the whole lymph node except some residual lymphoid follicles. This pattern suggests a continuous spread and growth of ECGs in sarcoid reactions along T-zone, where antigen presentation mainly takes place. Irrespective of granuloma type, a constant invasion of freshly recruited MRP8 + and MRP14 + macrophages was observed. Similar to sarcoidosis, angiotensin I-converting enzyme expression was a constant finding in epithelioid and giant cells, suggesting a common inflammatory pathway. An increasing ratio of CD4 + to CD8 + T lymphocytes (r = 0.789, P = .001) and a decreasing number of S100 + and CD83 + dendritic cells (r = 0.787, P = .001) within ECGs correlated with granuloma growth, whereas CD1a + immature dendritic cells were never observed inside ECGs. Our findings show that sarcoid reactions represent a T-cell-mediated immune response, leading to histological appearance and cell distribution similar to sarcoidosis and other granulomatous conditions, but the mechanism is different from dendritic cell-based tumor vaccination. Furthermore, mature dendritic cells occur inside ECGs especially of early sarcoid reactions but may not be required for the enlargement and further maintenance of ECGs, in contrast to CD4 + lymphocytes.

Adult↗

PubMed2004

Systemic granulomatous necrotizing vasculitis in a MPO-ANCA-positive patient.

We present a case of myeloperoxidase antineutrophil cytoplasmic antibody (MPO-ANCA)-associated vasculitis that demonstrated a systemic granulomatous lesion at autopsy. The patient initially showed anorexia, general malaise and anemia. Colon fiber was examined to detect the bleeding site, which revealed ischemic mucosal damage associated with venous fibrin thrombus. Because a high titer of MPO-ANCA was found, ANCA-associated vasculitis was suspected and the patient was started on steroid pulse therapy. However, anemia, renal failure and respiratory failure worsened and the patient died of sudden cardiac failure 2 days after the start of the therapy. An autopsy revealed systemic arteritis in multiple organs including the kidneys, liver, spleen, gastrointestinal system and genital organs that indicated fibrinoid necrosis accompanied by granulomatous reaction with multinucleated giant cells; the granulomatous reaction further extended along the splenic capsule. Glomerulonephritis and diffuse pulmonary damage, which are common in MPO-ANCA-associated vasculitis, were almost absent but parapleural fibrosis was present. The direct cause of death was presumed to be hemorrhagic shock due to rupture of an aneurysm in the gastric subserosa. As far as we know, this is the first case of a systemic granulomatous reaction in MPO-ANCA-positive vasculitis, although the cause of the granulomatous lesion is unknown.

Aged↗

PubMed2003

Multiple ulcers with perforation of the small intestine in buerger's disease: a case report.

We present the case of a young man with Buerger's disease that involved the gastrointestinal tract. The patient experienced sudden onset of abdominal pain during the period of treatment for peripheral manifestations. Radiographic workup revealed free air in the abdomen. A laparotomy was performed, and the resected specimen revealed regularly distanced and multiple ulcers in proximity of the perforation, which was accompanied by occluded intramural arteries of the small intestine. Although visceral localization of Buerger's disease is unusual, several cases have been previously reported. Nevertheless, the manifestation of regularly distanced skip intestinal ulcers in Buerger's disease is apparently unique. In addition, detailed pathological findings delineated this rare condition.

Adult↗

PubMed2003

Different immunophenotypes in Buerger's disease.

Recently, we reviewed the morphology of 31 specimens of thromboangiitis obliterans (TAO, Buerger's disease) in a multivariate analysis and showed that certain novel features of the affected vessels are different from arteriosclerosis obliterans (ASO) and thromboembolism. However, the pathogenic concept of TAO is still controversial. We applied immunohistochemistry to 58 amputated lower extremities and five autopsy controls. At specific sites of the diseased vessels, different cellular components were immunotyped by CD3, CD4, CD20, CD31, CD68, actin and desmin. These results were carefully compared among different diagnostic groups of vasoocclusive lesions by statistical methods. Some unique characteristics of TAO were identified when compared with ASO or thromboembolism. Consistent with a primary inflammatory and immunogenic lesion, lymphocytes and especially CD4+ T cells emerged significantly in TAO vessels and their adventitia. In the subset of definite TAO cases defined by all clinical criteria, the linear arrangement of macrophages, and B- and T-lymphocytes along vascular elastic fibers was the most striking additional finding, suggesting elastic fibers are an important immunogen. However, this feature was not apparent in closely related cases, otherwise similar to TAO and different from ASO and thromboembolism. Thus, our results indicate a heterogeneous group of TAO diseases, suggesting that damage to elastic fibers may be a secondary change to primary inflammation.

Actins↗