Binding characteristics of IgA 16.4.12E, a monoclonal antibody with specificity for the nonreducing terminal epitope of alpha-(1----6)-dextrans. Comparisons between IgA hybridoma 16.4.12E and myeloma W3129.

IgA 16.4.12E is a murine monoclonal antibody obtained following immunization with isomaltohexose linked to keyhole limpet hemocyanin. We have studied its interaction with methyl alpha-D-glucopyranoside and its derivatives bearing deoxy or deoxyfluoro groups, and with the methyl alpha-glycosides of a series of isomalto-oligosaccharides, some bearing deoxy or deoxy-fluoro groups at selected positions. From the data it is concluded that the antibody binds optimally to 4 sequential glucopyranosyl residues and that the protein subsite possessing the major affinity binds the terminal, nonreducing glucosyl group of that antigenic epitope. All the hydroxyl groups of that terminal glucosyl group are involved in hydrogen bonding, some in a donating and some in an accepting capacity. In the last part of the paper we report the construction of a possible model of the antibody, derived from its known amino acid sequence and the known crystalline structures of two closely related antibodies. It shows a pronounced cavity in the general immunoglobulin combining area which is flanked by 2 solvent-exposed tryptophanyl residues. A model recently reported for anti-dextran IgA W3129 shows a similar cavity with one such residue. Guided by hydrogen bonds, experimentally deduced from the comparison of the affinities of variously derivatized ligands, we suggest a speculative fitting for the nonreducing terminus of the dextran antigen, in the respective cavities of both IgA 16.4.12E and W3129.     

Influence of nitrogen/carbon ratio and complementary sugars on dextransucrase production by Leuconostoc mesenteroides NRRL B512(f).

The influence of the nitrogen/carbon ratio was studied during fermentation with Leuconostoc mesenteroides NRRL B512(f) using sucrose as substrate. The enzyme concentration was measured through activity tests and radiotracer tests with [¹⁴C]phenylalanine. In batch fermentation a slowing down in the rate of enzyme synthesis was observed with a decrease in the nitrogen/carbon ratio. Addition of pulses of nitrogen when the nitrogen/carbon ratio decreased allowed a constant production rate of enzyme and lower fermentation time. The influence of complementary sugars was also addressed. Lactose inhibited enzyme production. When galactose was used the yield was the same as in the fermentation with sucrose alone, but with a different production rate. Maltose favoured the synthesis of dextransucrase.     

Effects of apoA-V on HDL and VLDL metabolism in APOC3 transgenic mice

Apolipoprotein A-V (apoA-V) and apoC-III are exchangeable constituents of VLDL and HDL. ApoA-V counteracts the effect of apoC-III on triglyceride (TG) metabolism with poorly defined mechanisms. To better understand the effects of apoA-V on TG and cholesterol metabolism, we delivered apoA-V cDNA into livers of hypertriglyceridemic APOC3 transgenic mice by adenovirus-mediated gene transfer. In response to hepatic apoA-V production, plasma TG levels were reduced significantly as a result of enhanced VLDL catabolism without alternations in VLDL production. This effect was associated with reduced apoC-III content in VLDL. Increased apoA-V production also resulted in decreased apoC-III and increased apoA-I content in HDL. Furthermore, apoA-V-enriched HDL was associated with enhanced LCAT activity and increased cholesterol efflux. This effect, along with apoE enrichment in HDL, contributed to HDL core expansion and alpha-HDL formation, accounting for significant increases in both the number and size of HDL particles. As a result, apoA-V-treated APOC3 transgenic mice exhibited decreased VLDL-cholesterol and increased HDL-cholesterol levels. ApoA-V-mediated reduction of apoC-III content in VLDL represents an important mechanism by which apoA-V acts to ameliorate hypertriglyceridemia in adult APOC3 transgenic mice. In addition, increased apoA-V levels accounted for cholesterol redistribution from VLDL to larger HDL particles. These data suggest that in addition to its TG-lowering effect, apoA-V plays a significant role in modulating HDL maturation and cholesterol metabolism.     

Structure and evolution of <Emphasis Type="Italic">Apetala3</Emphasis>, a sex-linked gene in <Emphasis Type="Italic">Silene latifolia</Emphasis>

Background  

The evolution of sex chromosomes is often accompanied by gene or chromosome rearrangements. Recently, the gene AP3 was characterized in the dioecious plant species Silene latifolia. It was suggested that this gene had been transferred from an autosome to the Y chromosome.     

A new species of <Emphasis Type="Italic">Cystoisospora</Emphasis> Frenkel, 1977 (Apicomplexa: Sarcocystidae) from <Emphasis Type="Italic">Oecomys mamorae</Emphasis> Thomas (Rodentia: Cricetidae) in the Brazilian Pantanal

Despite the great diversity of coccidians, to our knowledge, no coccidian infections have been described in Oecomys spp. In this context, we examined Oecomys mamorae Thomas (Rodentia: Cricetidae) from the Brazilian Pantanal for infections with enteric coccidia. Nine individuals were sampled, and one was found to be infected. The oöcysts were recovered through centrifugal flotation in sugar solution. Using morphological and morphometric features, we described a new species of Cystoisospora Frenkel, 1977. Sporulated oöcysts were ovoidal 20.0–29.1 × 16.4–23.2 (26.7 × 21.2) µm and contained two sporocysts, 12.9–19.1 × 9.4–13.9 (16.4 × 12.4) µm, each with four banana-shaped sporozoites. Polar granule and oöcyst residuum were both absent. We documented the developmental forms in the small intestine and described the histopathological lesions in the enteric tract. Our results indicate that the prevalence of Cystoisospora mamorae n. sp. in O. mamorae is low, and tissue damage in the enteric tract is mild, even in the presence of coccidian developmental stages.     

Ultrastructure of the sodium pump: Comparison of thin sectioning, negative staining, and freeze-fracture of purified, membrane-bound (Na+, K+)-ATPase

Purified (Na+, K+)-ATPase was studied by electron microscopy after thin sectioning, negative staining, and freeze-fracturing, particular emphasis being paid to the dimensions and frequencies of substructures in the membranes. Ultrathin sections show exclusively flat or cup-shaped membrane fragments which are triple-layered along much of their length and have diameters of 0.1-0.6 μm. Negative staining revealed a distinct substructure of particles with diameters between 30 and 50 A and with a frequency of 12,500 +/- 2,400 (SD) per μm(2). Comparisons with sizes of the protein components suggest that each surface particle contains as its major component one large catalytic chain with mol wt close to 100,000 and that two surface particles unite to form the unit of (Na+,K+)-ATPase which binds one molecule of ATP or ouabain. The further observations that the surface particles protrude from the membrane surface and are observed on both membrane surfaces in different patterns and degrees of clustering suggest that protein units span the membrane and are capable of lateral mobility. Freeze-fracturing shows intramembranous particles with diameters of 90-110 A and distributed on both concave and convex fracture faces with a frequency of 3,410 +/- 370 per μm(2) and 390 +/- 170 per μm(2), respectively. The larger diameters and three to fourfold smaller frequency of the intramembranous particles as compared to the surface particles seen after negative staining may reflect technical differences between methods, but it is more likely that the intramembranous particle is an oliogomer composed of two or even more of the protein units which form the surface particles.     

A single amino acid change (Y318F) in the L-arabitol dehydrogenase (LadA) from Aspergillus niger results in a significant increase in affinity for D-sorbitol

Background  

L-arabitol dehydrogenase (LAD) and xylitol dehydrogenase (XDH) are involved in the degradation of L-arabinose and D-xylose, which are among the most abundant monosaccharides on earth. Previous data demonstrated that LAD and XDH not only differ in the activity on their biological substrate, but also that only XDH has significant activity on D-sorbitol and may therefore be more closely related to D-sorbitol dehydrogenases (SDH). In this study we aimed to identify residues involved in the difference in substrate specificity.     

Rapid Development and Optimization of Tablet Manufacturing Using Statistical Tools

The purpose of this paper was to develop a statistical methodology to optimize tablet manufacturing considering drug chemical and physical properties applying a crossed experimental design. The assessed model drug was dried ferrous sulphate and the variables were the hardness and the relative proportions of three excipients, binder, filler and disintegrant. Granule properties were modeled as a function of excipient proportions and tablet parameters were defined by the excipient proportion and hardness. The desirability function was applied to achieve optimal values for excipient proportions and hardness. In conclusion, crossed experimental design using hardness as the only process variable is an efficient strategy to quickly determine the optimal design process for tablet manufacturing. This method can be applied for any tablet manufacturing method.