Jun-Ichiro Murayama

Primary structure of horse erythrocyte glycophorin HA. its amino acid sequence has a unique homology with those of human and porcine erythrocyte glycophorins

SummaryThe complete amino acid sequence of the major sialoglycoproteins of horse erythrocyte membranes, glycophorin HA, was determined by manual sequencing methods, using tryptic, chymotryptic, and cyanogen bromide fragments. Glycophorin HA is a polypeptide chain of 120 amino acid residues and contains 10 oligosaccharide units attached to the amino-terminal side of the molecule. Its amino terminus is pyroglutamic acid. All of the oligosaccharides are linked O-glycosidically to threonine or serine residues. The amino acid sequence is consistent with the transmembrane orientation of glycophorins.There is no significant homology between the glycosylated domains of horse, human, and porcine glycophorins, but there is a considerable homology between the hydrophobic domains of the three glycophorins, which interact with the lipid bilayer of the erythrocyte membrane.

Fluorimetric assay of sialic acids

Abstract A fluorimetric assay has been developed for sialic acids in which sialic acids react with pyridoxamine to give fluorescent compounds in the presence of zinc ion and pyridine. This assay method is specific for unbound sialic acids and is a simple and sensitive procedure compared with the thiobarbituric acid assay of sialic acids.

Amino acid sequence and oligosaccharide attachment sites of the glycosylated domain of dog erythrocyte glycophorin

Glycophorin was prepared from dog erythrocyte membranes by extraction with lithium diiodosalicylate and partition in aqueous phenol. Tryptic and chymotryptic treatments of the glycophorin produced two major glycopeptides labeled T1 and CH1, respectively. The glycopeptides were isolated by gel chromatography followed by ion-exchange chromatography, and subjected to amino acid sequence analysis. Both glycopeptides represented the amino-terminal domain of the major dog glycophorin; T1 of 52 residues and CH1 of 43 residues. The amino-terminal sequence of dog glycophorin does not have significant homology with those of human, horse or porcine glycophorins. This result is in good agreement with our previous proposal that there is no homology in the sequence of the amino-terminal glycosylated domain of glycophorin.

Amino acid sequence of monkey erythrocyte glycophorin MK. Its amino acid sequence has a striking homology with that of human glycophorin A

A major sialoglycoprotein, glycophorin MK, was isolated from monkey erythrocyte membranes by extraction with lithium diiodosalicylate and partition in aqueous phenol. Chemical analysis of glycophorin MK revealed that the glycophorin consists of 51% protein and 49% carbohydrate by weight, and contains no N-glycosidic oligosaccharide units. Only N-glycolylneuraminic acid (Neu5Gc) was detected as sialic acid. The amino acid sequence of glycophorin MK was determined, which demonstrated that the glycophorin consists of 144 amino acid residues and 18 oligosaccharide units linked O-glycosidically to the peptide backbone through serine or threonine residues. The molecular weight was estimated to be about 35,000 based on the amino acid residues and carbohydrate content. By comparison of the amino acid sequence with those of human, equine and porcine glycophorins, a striking sequence homology was observed between monkey and human glycophorin. Glycophorin MK demonstrated both M and N blood group activities.

Structural studies of O-glycosidic oligosaccharide units of dog erythrocyte glycophorin

Abstract Dog glycophorin, the major sialoglycoprotein of dog erythrocyte membranes, contains either N -acetyl- or N -glycolylneuraminic acid, depending upon the strain of dog. Glycolipids also contained the same sialic acid as those found in glycophorin when both materials are prepared from erythrocyte membranes of individual dogs. The O -glycosidic oligosaccharides were released from glycophorin, prepared from individual dogs, by alkaline borohydride treatment, and purified by gel filtration and ion-exchange chromatography. The structures of the reduced oligosaccharides were determined by methylation analysis and gas-liquid chromatography-mass spectrometry. The O -glycosidic oligoscharides identified were one tetrasaccharide - Neu5Ac(2→3)Gal)1→3)[Neu5Ac(2→6)[GalNAcol - two trisaccharides - Neu5Ac(2→3)Gal1→3)GaINAcol and Gal(1→3)[Neu5Ac(2→6)]GalNAcol.

Development of a novel fluorometric assay for nitric oxide utilizing sesamol and its application to analysis of nitric oxide‐releasing drugs

Nitric oxide (NO) is related to various physiological effects as well as to numerous diseases caused by accentuation of NO production. Measurement of NO in cells and tissues is difficult as NO readily reacts with other molecules; furthermore, its half-life as a radical is fleeting. Currently, many NO pharmaceuticals are marketed as therapeutic agents for ischemic disease. Consequently, the identification of NO radicals and determination of generation rate from pharmaceuticals is very important when the effect of the medicinal supply is estimated. In this study, we developed a fluorometric assay for NO employing sesamol (3,4-methylenedioxyphenol) as a fluorometric substrate. Sesamol is converted to a fluorescent derivative (ex. 365 nm, em. 447 nm), which is dimmer in the presence of NO. The detection limit of NO with this method is 400 fmol; moreover, NO generated from drugs can be measured.

Structure of the majorO-glycosidic oligosaccharide of monkey erythrocyte glycophorin

Sialic acids and the majorO-glycosidic oligosaccharide of glycophorin MK from monkey (Japanese monkey,Macaca fuscata) erythrocyte membranes were characterized.N-Glycolylneuraminic acid (neu5Gc) was found as the major sialic acid, which was confirmed by gas-liquid chromatography-mass spectrometry as the trimethylsilyl methyl ester. ThreeO-glycosidic oligosaccharide units were obtained from a tryptic glycopeptide that contained all of the carbohydrate units in glycophorin MK by mild alkaline borohydride/borotritide treatment. Carbohydrate analyses of the oligosaccharides revealed that they were composed of Neu5Gc, galactose andN-acetylgalactosaminitol in the molar ratios of 1∶1∶1 (trisaccharide), 2∶1∶1 (tetrasaccharide) and 1∶1∶1 (pentasaccharide). The content of oligosaccharide units was estimated to be 1∶12∶5 for penta-, tetra- and trisaccharide, respectively, based on the yields, the molecular weight, and the number of oligosaccharide attachment sites in the amino-acid sequence. The tetrasaccharide was the major oligosaccharide and its structure was proposed to be Neu5Gcα2-3Galβ1-3[Neu5Gcα2-6]GalNAcol.

Development and evaluation of a formula for predicting introduction of medication self-management in stroke patients in the Kaifukuki rehabilitation ward

BackgroundMedication self-management in stroke patients is important to prevent further progression of disease and incidence of side effects. The purpose of this study was to create a formula for predicting medication self-management introduction in stroke patients using functional independence measure items and patient data, including medication-related information.MethodsThis was a retrospective analysis of 104 patients (cerebral infarction, cerebral hemorrhage, subarachnoid hemorrhage) discharged from the Kaifukuki rehabilitation ward at Showa University Fujigaoka Rehabilitation Hospital from January to December 2012. Multivariate analysis was performed to develop a formula for predicting achievement of medication self-management.ResultsOf the 104 patients, 39 (37.5%) achieved medication self-management. In the logistic regression analysis, number of drugs, age, walk/wheelchair mobility FIM, and memory FIM were extracted as significant factors independently contributing to achievement of medication self-management (p < 0.05). The prediction formula was [4.404 − 0.229 × number of drugs at admission + 0.470 × walk/wheelchair mobility FIM at admission + 0.416 × memory FIM at admission − 0.112 × age].ConclusionsIn the future, this formula may be used as an index to predict success of medication self-management in stroke patients.

Expected Duties of Pharmacists and Potential Needs of Physicians and Nurses on a Kaifukuki Rehabilitation Ward

This study investigated the required duties of pharmacists in a kaifukuki rehabilitation ward from the viewpoint of the ward physicians and nurses. A questionnaire survey was distributed to 27 facilities with kaifukuki rehabilitation wards. The questionnaire examined which duties the physicians and nurses expected from pharmacists while on the ward (4 areas, 10 items), as well as the time required for pharmacists to carry out those duties. Multivariate analysis was used to investigate which types of work took the most time for pharmacists on kaifukuki rehabilitation wards. Responses were received from 43 physicians and 184 nurses who worked on the kaifukuki rehabilitation wards of 19 facilities. The results revealed that the essential duties performed by pharmacists were the management of medical supplies, instruction on the use of self-medicating drugs at the time of introduction, and monitoring drug side effects. Furthermore, some duties, such as the distribution of medicines and changing or suggesting new drugs, required pharmacists to spend extended time on the ward. The responses indicated that physicians and nurses recognized the necessity for pharmacists to perform ward duties along with their routine work. This study shows that physicians and nurses working in kaifukuki rehabilitation wards demand proactive participation from pharmacists in appropriate medical therapy, such as instruction in the administration of medications and assessment at the time of prescription changes.

[Effect of the active adult learning/patient oriented clerkship on affective reaction of students ∼ from the results of student survey].

We have previously reported the efficacy of the Patient Oriented Clerkship (POC) in the clinical clerkship in Showa University Hospitals, by a trial with old four-year pharmacy program students. In the unique clerkship, each student has a patient in charge, and follows his/her clinical conditions throughout the rotation. The aim of the POC is that having the students learn spontaneously (Active Learning) and actively (Adult Learning) promoted by students commitment and responsibility by communicating with patients and health professionals in a team. As the POC requires students both Active Learning and Adult Learning, we define the POC as Active Adult Learning (AAL). Having a patient in charge for each student gives them many opportunities to participate in the medical team and foster their problem solving skills. Our previous study eventually showed positive results of the POC in the one-month short clerkship in the four-year program. On the other hand, the effect of the unique hospital clerkship in the new six-year program is not known. We conducted a student survey to clarify the learning effect in the new six-year education system which was revised and 2.5 month clinical clerkship was scheduled according to the model core clerkship curriculum. This report is the first report to show a challenge of the AAL/POC clerkship in the new six-year pharmacy education program.