Pelvic Artery Calcification Score Is a Marker of Vascular Calcification in Male Hemodialysis Patients.

Patients who undergo hemodialysis often suffer from cardiovascular disease (CVD), and evaluation of coronary artery calcification is extremely important. These evaluations are typically conducted using a noninvasive method including electron beam computed tomography (CT) or multi-detector CT, and the Agatston method to calculate the coronary artery calcification score (CACS). However, it is difficult to use for patients undergoing dialysis. Because patients undergoing dialysis is too strong in coronary artery calcification, and results become incorrect. Therefore, we were looking for a calcified evaluation place peculiar to a patients undergoing dialysis. We obtained pelvic artery calcification scores (PACS) using a 64-row multi-slice CT to assess the presence of calcification within a triangular space bordered by bordered by osseous structure. We used the Agatston method to calculate PACS. We compared male patients undergoing dialysis with male patients with normal renal function. Patients undergoing hemodialysis had a significantly higher incidence of pelvic artery calcification than normal controls (79.7% vs. 5.5%). In the dialysis group, CACS was 1660.2 (0-9056.1), and PACS was 48.8 (0-2943.1). We found a correlation between PACS and CACS and between PACS and dialysis period. We found penile artery calcification in male patients undergoing hemodialysis was more than normal controls, and it was possible to quantify PACS using the Agatston method. This study suggested the possibility that PACS became the vascular calcification evaluation method of the hemodialysis patient.

Synthesis of sugar fatty acid esters using a ß-xylosidase from Aspergillus awamori for transxylosylation.

Aspergillus awamori K4 ß-xylosidase has broad acceptor specificity. It has been used to synthesize a sugar fatty acid ester via its transxylosylation activity. One xylosyl residue was initially transferred to hexamethylene glycol as a linker with a yield of 0.36 g/g xylobiose. Linoleic acid was subsequently linked to one terminal hydroxyl side of the transfer product hydroxyhexyl xyloside through an esterification reaction catalyzed by a lipase. The synthesis of hexyl linoleoyl xyloside was confirmed by TOF-MS analysis. The binding with a linker improved the esterification reaction because of the hydrophobic hexamethylene chain and also prevented steric hindrance by the xylosyl residue. This sugar fatty acid ester synthesis method using transglycosylation should facilitate the production of emulsifiers or surfactants with various functions.

Production of L-arabinose from corn hull arabinoxylan by Arthrobacter aurescens MK5 α-L-arabinofuranosidase.

Arabinoxylans, which are comprised of a xylan backbone to which are attached glycosyl units that are primarily L-arabinofuranosyl units, are ubiquitous among plant species where it is a constituent of the cell wall. Arabinoxylan has attracted much attention as a potential biomass resource and L-arabinose has recently been reported to possess functional properties that are effective in the treatment of diabetes. Here, we report an α-L-arabinofuranohydrolase, isolated from the soil microbe Arthrobacter aurescens strain MK5, effective in releasing L-arabinose from corn hull arabinoxylan. When A. aurescens strain MK5 was grown in a liquid medium, corn hull arabinoxylan, which has a higher arabinose content (Ara/Xyl = 0.6) than oat spelts xylan (Ara/Xyl = 0.12), induced more efficient arabinoxylan hydrolase production. Analysis of enzyme activity in the culture broth revealed that arabinoxylan hydrolase activity was high, and α-L-arabinofuranosidase and ß-xylosidase activities were low. The optimum pH of the MK5 arabinoxylan hydrolase at 40 °C was around 7 and enzyme activity was relatively stable at an alkaline pH up to 9.5. The optimum temperature at pH 7 was around 50 °C and enzyme activity was stable under 50 °C. During the hydrolysis of corn hull arabinoxylan, only L-arabinose was released and 45.1% maximum sugar recovery was achieved. The A. aurescens MK5 enzyme was a typical arabinoxylan α-L-arabinofuranohydrolase and was most effective at releasing L-arabinose from corn hull arabinoxylan, which has a high arabinose content. This enzyme may have important industrial applications.

Production of fructooligosaccharides by beta-fructofuranosidases from Aspergillus oryzae KB.

Aspergillus oryzae KB produces two types of beta-fructofuranosidases: F1 and F2. F1 produces the fructooligosaccharides (FOSs) 1-kestose, nystose, and fructosyl nystose from sucrose through a transfructosylation action, whereas F2 mainly hydrolyzes sucrose to glucose and fructose. F1 and F2 enzymes were more selectively produced from the KB strain in liquid media with a sucrose concentration>2% and <2%, respectively. Immobilization using an anion-exchange resin (WA-30; polystyrene with tertiary amine) and cross-linking with glutaraldehyde depressed the hydrolysis reaction of F2 (high hydrolyzing enzyme) alone and enhanced the thermal stability of F1 (high transferring enzyme). F1 enzyme produced in the high sucrose medium was immobilized, cross-linked, and packed in a tubular reactor for continuous production of FOSs (24.6% 1-kestose, 21.6% nystose, 5.7% and fructosyl nystose). In a long-term operation in which 60% sucrose was imputed at 55 degrees C, the composition of FOSs produced was 51.9% (transfer ratio: 92%), and production by the immobilized enzyme was maintained for 984 h.

Two types of beta-fructofuranosidases from Aspergillus oryzae KB.

Aspergillus oryzae KB produces two types of beta-fructofuranosidases, F1 and F2. F1 produces 1-kestose, nystose, and fructosyl nystose from sucrose through its transfructosylation action. F2 hydrolyzes sucrose to glucose and fructose. N-Terminal amino acid sequences of the purified enzymes were DYNAAPPNLST for F1 and YSGDLRPQ for F2. Each enzyme encoding gene was identified in the genome of Aspergillus oryzae. Although the KB strain showed a higher production of F2 than F1 in a low sucrose liquid medium, F2 production gradually decreased, whereas F1 production increased with increasing sucrose concentration in the medium. Synthesis of F1 and F2 mRNAs analyzed on reverse-transcription polymerase chain reaction corresponded to individual enzymatic production. During liquid culture of the KB strain, F1 synthesizes fructooligosaccharides from sucrose through transfructosylation, and F2 gradually hydrolyzes it. In a highly concentrated sucrose medium, intake of sucrose into the KB strain was depressed by F1 through synthesis of transfer products, fructooligosaccharides.

Biological pretreatment with two bacterial strains for enzymatic hydrolysis of office paper.

The cellulose-hydrolyzing strains, Sphingomonas paucimobilis MK1 and Bacillus circulans MK2, were separated from soil and were grown together in a single culture plate. Growth B. circulans MK2 in liquid culture required symbiosis with S. paucimobilis MK1. Biological pretreatment with the combined strain suspension after the liquid culture improved enzymatic hydrolysis of office paper from municipal wastes. Sugar recovery by S. paucimobilis MK1 (51%) was 1.4 times higher than that of the untreated sample (30%) and in the strain combination with B. circulans MK2, recovery was further improved by 2.5 times (75%). The sugar recovery in maximum condition was enhanced up to 94% for office paper. Furthermore, biological pretreatment effects were confirmed for more than 1 day less time. In X-ray diffraction patterns for the crystallinity of cellulose in office paper changed after biological pretreatment, the crystallinity was increased in comparison to that in untreated paper. The mechanism of biological pretreatment effect was explained by the fact that the strain acted as an endoglucanase, which hydrolyzes amorphous areas randomly.

Production of galacto-manno-oligosaccharides from guar gum by beta-mannanase from Penicillium oxalicum SO.

Beta-mannanase from Penicillium oxalicum SO efficiently hydrolyzed guar galactomannan to galacto-manno-oligosaccharides. Gel filtration estimated the molecular weight of the beta-mannanase as 35 000 and SDS-PAGE as 29 000. The optimum pH was around 5 while a stable pH was reached in the range of 3-6. Optimum temperature was around 60 degrees C at pH 5, while under 60 degrees C activity was stable. HPLC analysis detected oligosaccharides with degrees of polymerization (DP) of 2 to 7 and 2 to 6 released on hydrolysis of guar and locust bean gums, respectively; about 92% of the released sugars were oligosaccharides. In analysis of the sugar distribution on MALDI-TOF-MS, major products of DP 6 and 7 and DP 5 and 6 were confirmed in hydrolysates of guar gum and locust bean gum, respectively. One of the main oligosaccharides released from guar gum, with DP 7, had a high galactose content (Gal/Man = 0.76) and corresponded to a blockwise galactose-substituted mannan type in galactomannan.

Characteristics of transxylosylation by beta-xylosidase from Aspergillus awamori K4.

The Aspergillus awamori K4 beta-xylosidase gene (Xaw1) sequence was deduced by sequencing RT-PCR and PCR products. The ORF was 2,412 bp and the predicted peptide was 804 amino acids long, corresponding to a molecular weight of 87,156 Da. The mature protein was 778 amino acids long with a molecular weight of 84,632 Da. A homology search of the amino acid sequence revealed that it was very similar to the Aspergillus niger beta-xylosidase gene with only five amino acid differences. K4 beta-xylosidase had the same catalytic mechanism as family 3 beta-glucosidases, involving Asp in region A. At an early stage in the reaction with xylobiose and xylotriose, the hydrolysis rate was much lower than the transxylosylation rate, decreasing gradually as the substrate concentration increased, whereas the transxylosylation rate increased greatly. Aspergillus awamori K4 beta-xylosidase had broad acceptor specificity toward alcohols, hydroxybenzenealcohols, sugar alcohols and disaccharides. A consensus portion involving the hydroxymethyl group of the acceptor was confirmed in the major transfer products 1(4)-O-beta-D-xylosyl erythritol, (2-hydroxyl)-phenyl-methyl-beta-D-xylopyranoside, 6S-O-beta-D-xylosyl maltitol (S: sorbitol residue) and 6G-O-beta-D-xylosyl palatinose (G: glucosyl residue). This might suggest that the methylene in the hydroxymethyl group facilitates base-catalyzed hydroxyl group attack of the anomeric center of the xylosyl-enzyme intermediate.

Synthesis of new glycosides by transglycosylation of N-acetylhexosaminidase from Serratia marcescens YS-1.

Serratia marcescens YS-1, a chitin-degrading microorganism, produced mainly N-acetylhexosaminidase. The purified enzyme had an optimal pH of approximately 8-9 and remained stable at 40 degrees C for 60 min at pH 6-8. The optimum temperature was around 50 degrees C, and enzyme activity was relatively stable below 50 degrees C. YS-1 N-acetylhexosaminidase hydrolyzed p-nitrophenyl beta-N-acetylgalactosamide by 28.1% relative to p-nitrophenyl beta-N-acetylglucosamide. The N-acetylchitooligosaccharides were hydrolyzed more rapidly, but the cellobiose and chitobiose of disaccharides that had the same beta-1,4 glycosidic bond as di-N-acetylchitobiose were not hydrolyzed. YS-1 N-acetylhexosaminidase efficiently transferred the N-acetylglucosamine residue from di-N-acetylchitobiose (substrate) to alcohols (acceptor). The ratio of transfer to methanol increased to 86% in a reaction with 32% methanol. N-Acetylglucosamine was transferred to the hydroxyl group at C1 of monoalcohols. A dialcohol was used as an acceptor when the carbon number was more than 4 and a hydroxyl group existed on each of the two outside carbons. Sugar alcohols with hydroxyl groups in all carbon positions were not proper acceptors.