Biomedical subjects

H Kurihara

Publications and source records attributed to H Kurihara.

At least 271 records · Page 15Linked to original sources

PubMed1993

Thyroplasty type I with ceramic shim.

To improve hoarseness or misswallowing caused by unilateral recurrent laryngeal nerve paralysis, medialization of paralyzed vocal cord has frequently been performed. This method includes such techniques as injection method, insertion method, and arytenoid adduction, each presenting its merits and demerits. The insertion method which can be done while monitoring the patient's voice seems advantageous in that the technique is easy to perform and generally guarantees the voice improvement. Among insertion methods, Isshiki thyroplasty type I is the one most representative as well as popularized. However, since a silicone shim is inserted in this operation, it may be accompanied by the risks of carcinogenicity, foreign body reaction, and induction of collagen disease of silicone. Therefore we planned to use a ceramic as a safe substitute instead of silicone. There has been no article reporting the use of ceramic in this type of surgery. We used a fibrin glue to fix the ceramic shim and for hemostasis, which was found very useful. Hitherto, 2 cases of unilateral recurrent laryngeal nerve paralysis underwent Isshiki thyroplasty type I using ceramic shim with satisfactory results.

Aged↗

PubMed1993

Association of Actinobacillus actinomycetemcomitans leukotoxin with nucleic acids on the bacterial cell surface.

Actinobacillus actinomycetemcomitans, a periodontopathic gram-negative bacterium, produces a leukotoxin that is a member of the RTX cytotoxin family. Although genes may function in toxin secretion, the leukotoxin is not secreted extracellularly but remains associated with the bacterial cell surface. We report here that this toxin-cell surface association is mediated by nucleic acids and directly demonstrate that the extracellular secretion of toxin occurs in growing cultures with increased ionic strength of medium. All examinations were performed with freshly harvested A. actinomycetemcomitans 301-b from anaerobic fructose-limited chemostat cultures. The occurrence of cell surface-localized DNA was shown by directly digesting whole cells with the restriction endonuclease EcoRI or HindIII, which yielded many DNA fragments. The cell surface DNA constituted about 20% of the total cellular DNA. The leukotoxin was released from the whole cells by digestion with DNase I as well as restriction endonucleases. Because the leukotoxin binds ionically to DNA, it is dependent on the ionic strength of buffers or media. Accordingly, the toxin was released from cells suspended in saline at pH 7.5 in the presence of increasing amounts of MgCl2 (0 to 10 mM) or NaCl (0 to 50 mM). Moreover, a considerable quantity of leukotoxin was detected in the culture supernatant of fructose-limited chemostat cultures when sodium succinate solution was pumped into the steady state as an additional salt (30 and then 50 mM). This toxin-DNA association was also found in well-characterized strains including not only the leukotoxin-producing ATCC 29522 but also the toxin production-variable ATCC 29523 and the non-leukotoxin-producing ATCC 33384 when these strains were grown in the chemostat culture.

Aggregatibacter actinomycetemcomitans↗

PubMed1993

Calcium-dependent protein kinase C activity of neutrophils in localized juvenile periodontitis.

Protein kinase C is a key molecule in neutrophil signal transduction after receptor stimulation by soluble bioactive molecules. It has been reported that neutrophils from most patients with localized juvenile periodontitis (LJP) do not have a normal response after stimulation with a chemotactic ligand such as N-formylmethionylleucylphenylalanine (FMLP). To further clarify the mechanism of this altered response and to confirm and expand earlier observations, the calcium-dependent protein kinase C activity of neutrophils from patients with LJP was evaluated. Peripheral blood neutrophils from 12 patients and 12 healthy subjects, age, sex, and race matched, were sonicated and subsequently subfractionated by ultracentrifugation into a soluble fraction (cytosol rich) and a particulate fraction (membrane rich). The calcium-dependent protein kinase C activity was evaluated in each fraction by phosphorylation of histone with radiolabeled ATP in the presence or in the absence of phorbol 12-myristate 13-acetate stimulation. Results revealed that the total calcium-dependent protein kinase C activity of neutrophils from patients with LJP and depressed chemotactic migration to FMLP (201.0 +/- 63.6 pmol/min/10(7) cells) was lower than that of neutrophils from healthy subjects (287.6 +/- 55.7 pmol/min/10(7) cells) (P < 0.01). The calcium-dependent protein kinase C activity in neutrophils from patients with LJP exhibited a positive correlation with chemotactic migration to FMLP (P < 0.05). The low activity of calcium-dependent protein kinase C in neutrophils from the patients reflected the low activity in the soluble fraction from the neutrophils. After stimulation with phorbol 12-myristate 13-acetate, the calcium-dependent protein kinase C activity was found to be lower from patients with LJP than from healthy subjects. These results suggest that lower calcium-dependent protein kinase C in neutrophils is a predisposing factor for LJP.

Aggressive Periodontitis↗

PubMed1993

Reperfusion after brief ischemia disrupts the microtubule network in canine hearts.

Histological changes in the stunned myocardium are believed to be minimal. This study examined whether cytoskeletal structures of microtubules are disrupted in the stunned myocardium. In 38 dogs, the left anterior descending coronary artery was occluded for 15 minutes and reperfused to produce the stunned myocardium. Microtubules were stained immunohistochemically. In intact myocardium, microtubules appeared as a filamentous network throughout the cytoplasm and encircled the nucleus. This pattern was not affected by 15 minutes of ischemia. One hour of reperfusion, however, disrupted microtubular structure substantially (disruption score in the endocardium, 53.4 +/- 6.0%) although actin filaments remained intact. Microtubular structures were reconstituted 1-3 days after reperfusion, showing supernormal immunoreactivity. Five days after reperfusion, the pattern of microtubular staining was normal. In another protocol, the role of Ca2+ during reperfusion in microtubular disruption was examined. When intracoronary infusion of EDTA (1.67 mumol/kg body wt per minute) was performed during the initial 10 minutes of reperfusion, myocardial stunning was attenuated. The fractional shortening in the perfused area after 1 hour of reperfusion was 20.1 +/- 1.2% versus 11.5 +/- 0.5% in the control condition (p < 0.05), and the microtubular disruption score was lower (12.6 +/- 1.4%). Although intracoronary infusion of calcium chloride (9 mumol/kg body wt per minute) for 10 minutes in nonischemic hearts increased contractile function (fractional shortening, 25.3 +/- 2.0%), it severely disrupted microtubular networks (microtubular disruption score, 64.0 +/- 10.6%). We conclude that microtubules supporting the structural integrity of myofibrils and other organelles are reversibly disrupted by reperfusion after brief ischemia probably through calcium overload.

Animals↗

PubMed1993

Disruption of cytoskeletal structures mediates shear stress-induced endothelin-1 gene expression in cultured porcine aortic endothelial cells.

Hemodynamic shear stress alters the architecture and functions of vascular endothelial cells. We have previously shown that the synthesis of endothelin-1 (ET-1) in endothelial cells is increased by exposure to shear stress. Here we examined whether shear stress-induced alterations in cytoskeletal structures are responsible for increases in ET-1 synthesis in cultured porcine aortic endothelial cells. Exposure of endothelial cells to 5 dyn/cm2 of low shear stress rapidly increased monomeric G-actin contents within 5 min without changing total actin contents. The ratio of G- to total actin, 54 +/- 0.8% in quiescent endothelial cells, increased to 87 +/- 4.2% at 6 h and then decreased. Following the disruption of filamentous (F)-actin into G-actin, ET-1 mRNA levels in endothelial cells also increased within 30 min and reached a peak at 6 h. The F-actin stabilizer, phalloidin, abolished shear stress-induced increases in ET-1 mRNA; however, it failed to inhibit increases in ET-1 mRNA secondary to other stimulants. This indicates that shear stress-induced increases in ET-1 mRNA levels may be mediated by the disruption of actin fibers. Furthermore, increases in ET-1 gene expression can be induced by actin-disrupting agents, cytochalasin B and D. Another cytoskeleton-disrupting agent, colchicine, which inhibits dimerization of tubulin, did not affect the basal level of ET-1 mRNA. However, colchicine completely inhibited shear stress- and cytochalasin B-induced increases in ET-1 mRNA levels. These results suggest that shear stress-induced ET-1 gene expression in endothelial cells is mediated by the disruption of actin cytoskeleton and this induction is dependent on the integrity of microtubules.

Actins↗

PubMed1993

Relationship between tissue polyamine levels and malignancy in primary brain tumors.

We studied the relationship between tissue polyamine levels and malignancy in 146 primary brain tumors. Astrocytoma showed a consistent rise in levels of N1-acetylspermidine (N1-AcSpd) in tissue with increased grade of malignancy. Furthermore, in astrocytoma, the patients whose tumors contained high levels of N1-AcSpd (> or = 15 nmol/g) in tissue showed a far worse prognosis, a significantly lower recurrence-free survival rate, and a shorter survival rate than did the patients whose tumors contained lower levels of N1-AcSpd (< 15 nmol/g). High-grade astrocytoma contained N1-AcSpd at the level of 15 nmol/g or higher, and most of the benign brain tumors, including low-grade astrocytoma, meningioma, pituitary adenoma, and normal brain tissues contained much lower levels of N1-AcSpd. However, medulloblastoma, hemangiopericytoma, and neurinoma showed an inverse correlation between N1-AcSpd levels in tissue and biological malignancy. These results suggest that levels of N1-AcSpd in tissue can be a promising biochemical marker of malignancy in astrocytoma. In the other primary brain tumors, however, the relationship between levels of N1-AcSpd in tissue and biological malignancy should be examined in each histopathological type.

Adolescent↗

PubMed1993

Abnormal proportion of gamma delta T cells in peripheral blood is frequently detected in patients with periodontal disease.

The proportions of gamma delta T cells in the peripheral blood of 8 early-onset periodontitis (EOP) patients, 8 later-onset periodontitis (LOP) patients, and of 17 healthy subjects (HS) were assessed by immunofluorescence assay using an anti-pan gamma delta T cell monoclonal antibody (TCR-delta 1) to study the population attributes of peripheral blood gamma delta T cells in patients with periodontal disease. The distribution of the proportions of gamma delta T cells in both EOP and LOP patients was found to be one and one half times broader than that in the HS; however, there was no significant difference in the proportion of gamma delta T cells between each group. An abnormally high proportion of peripheral blood gamma delta T cells in pan T cells (> 9.90%) was observed in 37.5% of both the EOP patients and the LOP patients. An abnormally low proportion (< 5.57%) was observed in 50% of the EOP patients and in 25% of the LOP patients. The higher occurrences of the high or low gamma delta T cell proportions were significant between EOP and HS, and between EOP+LOP and HS. There was no correlation between the proportion of gamma delta T cells and age or the clinical parameters of periodontal status. It was concluded that an abnormal proportion of gamma delta T cells is frequently detected in patients with periodontal disease. These findings are consistent with the proposed role of the gamma delta T cells for playing a unique role as a first line defense against infections at body surfaces.

Adolescent↗

PubMed1993

[A case of middle ear carcinoma with high plasma G-CSF level].

Malignant tumors sometimes yield a variety of paraneoplastic syndrome. We report a case of middle ear cancer demonstrating unusual granulocytosis which was considered to belong to this syndrome. A patient, 67-year-old male, noticed left facial nerve palsy in June 1989, and was operated on under the diagnosis of chronic otitis media. However, middle ear tumor was suspected during the operation, which was confirmed to be squamous cell carcinoma. He was referred to our hospital for radiotherapy. Six months after irradiation, spontaneous liqorrhea due to the dural necrosis occurred. When intracranial repair of dura was performed, tumor invasion into the dura was noticed. Subtotal resection of the temporal bone with iridium implant was done in vain to control the recurrent tumor, which spread quickly to adjacent tissues. A month before the death, progressive granulocytosis without appreciable inflammatory signs started. Numbers of WBC rapidly increased, maximally reaching 88,300/cmm. Plasma G-CSF concentration was assayed at this time, showing about ten times as much as that of normal controls. It was thus postulated that production of this substance by the tumor was responsible for granulocytosis.

Aged↗

PubMed1993

[The clinical significance of carbohydrate antigen 19-9 (CA19-9) in transitional cell cancer of urinary tract].

For the purpose of evaluating the utility of CA19-9 determination in transitional cell cancer (TCC) of the urinary tract, its serum value, tissue level and, immunohistochemical study, and follow up study, with high serum value were examined in 43 cases with TCC and 13 cases of control. 1. Serum values were 19.1 U/ml in the control group and 198 U/ml in the tumor group. Though those of tumor were higher than those of the control group, there is no statistical difference between both groups. The cases with abnormally high value were observed in 13.7% of the control group, 13.8% of the bladder cancer, and 57.1% of the upper urinary tract cancer. 2. Tissue levels were 816 U/g wet tissue in the control group and 13,000 U/g wet tissue in the tumor group, showing a significantly higher value (p < 0.01) in the tumor group, statistically. 3. Immunohistochemical study were positive in 4.0% of the control group and 88.4% of the tumor group. 4. There is a significant correlation between serum values and tissue levels (p < 0.05, r = 0.483). 5. The serum values did not correlated with the staining property, tumor size, grade, and stage. 6. The tissue levels correlated with the tumor size, but did not correlate with the staining property, grade, and stage. 7. In the follow-up study of cases with high value, those, who had not attained postoperative normalization, died of recurrent carcinoma. In cases showing an elevation despite postoperative normalization, too, all showed recurrence. Only those, whose value were normalized and remained at the normal range, are living without metastasis.(ABSTRACT TRUNCATED AT 250 WORDS)

Antigens, Tumor-Associated, Carbohydrate↗

PubMed1993

Relation of blood pressure circadian rhythm to cardiovascular complaints and orthostatic test results in patients with orthostatic dysregulation. Effects of midodrine on blood pressure circadian rhythm.

Twenty patients with orthostatic dysregulation were studied in order to clarify the relation of blood pressure (BP) circadian rhythm to cardiovascular complaints and orthostatic test results. There was a significant negative correlation between the number of cardiovascular complaints and 24 hour (hr) maximum (Mx) and day-time Mx BPd. A significant negative correlation was also obtained between the decrease in TII amplitude in upright posture and nighttime standard deviation and range of BPs and BPd. Decrease of BPs in upright posture was significantly correlated with 24 hr and nighttime minimum (Mn) BPd. The above results indicate that cardiovascular complaints and orthostatic test results are related with BPd, as well as BPs.

Blood Pressure↗

PubMed1993

Immunocytochemical and biochemical characterization of the Heymann nephritis antigenic complex in rat L2 yolk sac cells.

Heymann nephritis in the rat is the most widely used model of human membranous glomerulonephritis. Glycoprotein (gp)330, a large (M(r) > 550,000) membrane-associated glycoprotein, has been identified as the main antigen in this autoimmune disease. Studies of gp330 and receptor-associated protein (RAP), its 44-kd subunit, have been restricted largely to rat kidney, as no stable cultured cell line has been available that expresses gp330. We have recently identified a rat yolk sac carcinoma cell line (L2) that expresses both gp330 and RAP. In this report, we have carried out detailed morphological, immunocytochemical, and biochemical studies characterizing the biosynthesis and localization of gp330 and RAP in the L2 rat yolk sac cell line. At the electron microscope level, the L2 cells are seen to be attached by cell junctions, and their predominant morphological features include extensive networks of rough endoplasmic reticulum (ER) and numerous clathrin-coated pits found on the cell membrane. By immunocytochemistry, gp330 was localized primarily to clathrin-coated pits at the cell surface, whereas RAP was localized predominantly to the lumen of the rough ER. Pulse-chase experiments indicated that gp330 spends a prolonged time maturing in the ER of L2 cells, as transport of gp330 to the Golgi complex (based on acquisition of endoglycosidase H resistance) is slow (t1/2 = 90 to 120 minutes). Gp330 reached the L2 cell surface beginning at 2 hours after synthesis, where it could be detected by cell surface immunoprecipitation. RAP was found to be an N-linked glycoprotein, and it remained endoglycosidase H-sensitive up to 4 hours after synthesis. Co-precipitation and co-sedimentation experiments demonstrated that gp330 and RAP form a large heterodimer (M(r) approximately 669,000) immediately after biosynthesis and are further assembled into a large hetero-oligomer in the ER. These findings demonstrate that the localization and the kinetics of assembly of gp330 and RAP into the Heymann nephritis antigenic complex are similar in both L2 cells and rat kidney. They also provide new information on the intracellular processing of these two molecules and their delivery to the cell surface. Thus, the L2 cell system should facilitate further characterization of the functions and interactions of gp330 and RAP, which may shed light on the cellular and molecular mechanisms of Heymann nephritis.

Animals↗

PubMed1993

Human polymorphonuclear leukocytes have dual effects on endothelin-1: the induction of endothelin-1 mRNA expression in vascular endothelial cells and modification of the endothelin-1 molecule.

The effect of human polymorphonuclear leukocytes (PMNs) on the expression of the endothelin-1 (ET-1) gene and the production of ET-1 peptide was investigated. Human PMNs were separated from venous blood with Mono-Poly Resolving Medium and activated by incubation with formyl-methionyl-lencylphenylalanine (FMLP) (1 microM). Then PMN suspension was added to cultured porcine endothelial cell monolayers and coincubated for various periods. Following the coincubation, ET-1 mRNA in endothelial cells was examined by Northern blotting and immunoreactive ET-1 (irET-1) peptide levels in the conditioned media were measured by an enzyme-linked immunosorbent assay (ELISA). Similar experiments were also carried out with cell-free PMN supernatant. Untreated and activated PMNs led to a 1.4-fold and 6.3-fold increase in ET-1 mRNA levels in endothelial cells, respectively, at 6h, while irET-1 peptide levels did not significantly increase as compared with control. In contrast, when PMNs were coincubated in the presence of an Intercell chamber without direct contact to endothelial cells, PMNs did not induce ET-1 mRNA expression in endothelial cells, and significantly decreased irET-1 peptide levels in the conditioned media. Cell-free PMN supernatant did not have all these effects on ET-1. These findings suggest that direct PMN-endothelial cell contact was essential for PMN-induced expression of the ET-1 gene and that PMNs may decrease irET-1 through some modification of the ET-1 molecule.

Animals↗

PubMed1993

Effect of Ca2+ on the binding of Actinobacillus actinomycetemcomitans leukotoxin and the cytotoxicity to promyelocytic leukemia HL-60 cells.

The effects of Ca2+ on the binding of Actinobacillus actinomycetemcomitans leukotoxin (LT) to LT-susceptible promyelocytic leukemia HL-60 cells and the consequent cytotoxicity were investigated in comparison with those on LT-resistant erythroleukemia K-562 cells. The total amount of LT bound to the HL-60 and K-562 cells, as determined by ELISA using anti-LT rabbit serum, was essentially the same for both cells. The percentage of bound LT decreased in the presence of phosphatidylcholine (PC) was also almost equal for both cells. In contrast, EGTA added in the presence of PC enhanced the decrease of bound LT significantly from HL-60, but not from K-562. By monitoring the fluorescence of Fura-2 and of fluorescein as a measure for the change in intracellular calcium ions ([Ca2+]i) and cytotoxic activity, respectively, we showed that the exposure of HL-60 to LT induced the [Ca2+]i increase followed by the cell death, whereas this was distinctly not the case for K-562. These results indicate that a Ca(2+)-dependent LT-binding mechanism is possibly the step characteristic to LT-susceptible HL-60 cells.

Aggregatibacter actinomycetemcomitans↗

PubMed1993

[Case report of metastatic cutaneous angiosarcoma causing bilateral pneumothorax].

The patient was a 72-year-old man who underwent operation for bilateral pneumothorax due to rupture of cystic lung lesions. Two months after the operation, intrathoracic bleeding redeveloped. The patient developed a tumor on his head. Pathological examination of the scalp lesion revealed cutaneous angiosarcoma. A biopsy of thickened pleura showed that the bleeding was due to pleural metastatic angiosarcoma of the skin. The multiple cystic lesions were considered to be metastases of angiosarcoma. We conclude that pneumothorax or hemothorax in the elderly should be differentiated from malignant metastatic lung tumor. Cystic lung lesions should be considered as possible metastatic angiosarcoma from the skin.

Aged↗

PubMed1992

Multiple thyroid involvement (intraglandular metastasis) in papillary thyroid carcinoma. A clinicopathologic study of 105 consecutive patients.

Multiple thyroid involvement (MTI) in papillary thyroid carcinoma was clinicopathologically studied in 105 non-selected, consecutive patients. Whole thyroids resected by total thyroidectomy were sectioned at intervals of 2-3 mm (mean number of slices per gland, 19.2) and histologically reviewed. The intraglandular cancer foci, other than the tumor regarded as the primary focus, were demonstrated in 82 (78.1%) of 105 patients. The foci were usually small (less than 4 mm) and were not accompanied by sclerotic fibrous stroma or by a fibrous capsule. These small foci were distributed around the primary lesion and also were found frequently (61.0%) in the opposite lobe as bilateral disease. In the opposite lobe, a similar incidence (approximately 30%) of disease was obtained in each of the three parts (upper, middle, and lower). The mean number of foci in patients with MTI was statistically correlated to age, the presence of lymph node metastases, and the presence of solid areas or psammoma bodies in the primary tumor. It was concluded that MTI could be regarded as one of the most striking and important biologic characteristics of papillary thyroid carcinoma.

Adolescent↗

PubMed1992

gp330 associates with a 44-kDa protein in the rat kidney to form the Heymann nephritis antigenic complex.

Using antibodies isolated from glomeruli of nephritic rats we have previously identified a 330-kDa cell surface glycoprotein (gp330) as a major pathogenic antigen of Heymann nephritis (HN), an experimental model of human membranous glomerulonephritis. Recently, we have isolated a cDNA clone, C14, encoding a polypeptide that contains a pathogenic epitope of HN responsible for the initiation of the disease. Subsequently, another protein, alpha 2-macroglobulin receptor-associated protein (alpha 2-MRAP), which is a subunit of the receptor for human alpha 2-macroglobulin/low density lipoprotein receptor-related protein (LRP), was shown to possess a high degree of sequence homology to the C14 protein (C14p). In this report, we have investigated the relationship between gp330, C14p, and alpha 2-MRAP. Immunoprecipitation studies demonstrate that gp330 forms a heterodimeric association with a 44-kDa polypeptide that is stable to detergent extraction and long-term centrifugation. Further, immunoblotting analysis on the purified complex indicates that the 44-kDa associated protein shares immunological identity to C14p and alpha 2-MRAP. In addition, antibodies eluted from glomeruli of HN rats and antibodies to a C14 fusion protein immunoprecipitated gp330 and the 44-kDa protein, demonstrating that the epitopes responsible for the initial events of HN are accessible within the complex. Based on these data, three models are proposed to explain how pathogenic epitopes in the gp330-44-kDa, HN antigenic complex may be presented at the cell surface and initiate the onset of HN.

Animals↗

PubMed1992

Diversity among tight junctions in rat kidney: glomerular slit diaphragms and endothelial junctions express only one isoform of the tight junction protein ZO-1.

ZO-1 is a 225-kDa peripheral membrane protein present in all tight junctions. It was recently shown to consist of two isoforms that differ in the presence of an internal 80-amino acid domain termed motif-alpha. To obtain information on their distribution and potential functional significance we have localized the two isoforms in rat kidney by using antibodies that recognize either both ZO-1 isoforms or the larger, motif-alpha-containing isoform. By immunofluorescence, staining with both antibodies was demonstrated at all tight junctions of tubular epithelial cells and the epithelial cells of Bowman's capsule. In contrast, the motif-alpha-containing isoform was absent from the slit diaphragms of the glomerular epithelium and the tight junctions of glomerular and peritubular capillary endothelial cells. This restricted isoform expression was confirmed by immunoblot analysis comparing proteins from purified glomeruli with those from kidney cortex or medulla. Thus, while both isoforms are expressed in typical epithelial tight junctions, only a single isoform, lacking motif-alpha, is expressed in the highly specialized slit diaphragms, where the intercellular spaces are normally open, and in endothelial junctions, which are readily opened by physiologic signals. The differential expression of ZO-1 isoforms in structurally and functionally distinct junctions in the kidney suggests that they may contribute to defining the variable functional properties, in particular the lability of these intercellular junctions.

Aging↗

PubMed1992

Crystal and molecular structure of RNase Rh, a new class of microbial ribonuclease from Rhizopus niveus.

The crystal structure of RNase Rh, a new class of microbial ribonuclease from Rhizopus niveus, has been determined at 2.5 A resolution by the multiple isomorphous replacement method. The crystal structure was refined by simulated annealing with molecular dynamics. The current crystallographic R-factor is 0.200 in the 10-2.5 A resolution range. The molecular structure which is completely different from the known structures of RNase A and RNase T1 consists of six alpha-helices and seven beta-strands, belonging to the alpha+beta type structure. Two histidine and one glutamic acid residues which were predicted as the most probably functional residues by chemical modification studies are found to be clustered. The steric nature of the active site taken together with the relevant site-directed mutagenesis experiments (Irie et al.) indicates that: (i) the two histidine residues are the general acid and base; and (ii) an aspartic acid residue plays a role of recognizing adenine moiety of the substrate.

Amino Acid Sequence↗