Evidence for interaction between transmembrane segments in assembly of Kv1.3

Previously, we showed that the N-terminal recognition domain (T1) of Kv1.3 was not required for assembly of functional channels [Tu et al. (1996) J. Biol. Chem. 271, 18904-18911]. Moreover, specific Kv1.3 peptide fragments including regions of the central core are able to inhibit expression of current produced from a channel lacking the T1 domain, Kv1.3(T1-). To elucidate the mechanism whereby Kv1.3 peptide fragments suppress Kv1.3(T1-) current, we have studied the ability of peptide fragments containing the transmembrane segments S1, S1-S2, or S1-S2-S3 to physically associate with the Kv1.3(T1-) polypeptide subunit in vitro in microsomal membranes. Using c-myc (9E10) epitope-labeled peptide fragments and anti-myc antibody as well as antisera to the Kv1.3 C-terminus, we now demonstrate specific association of these peptide fragments with Kv1.3(T1-). Association of peptide fragments with Kv1.3(T1-) was correlated with integration of both proteins into the membrane. Furthermore, the relative strength and kinetics of this association directly correlated with the ability of fragments to suppress Kv1.3(T1-) current. The rate-limiting step in the sequential synthesis, integration, and formation of a complex was the association of integrated polypeptides within the plane of the lipid bilayer. These results strongly suggest that the physical association of transmembrane segments provides the basis for suppression of K+ channel function by K+ channel peptide fragments in vivo. Moreover, the S1-S2-S3 peptide fragment potently suppressed full-length Kv1.3, thus implicating a role for the S1-S2-S3 region of Kv1.3 in the assembly of the Kv1.3 channel. We refer to these putative association sites as IMA (intramembrane association) sites.     

Detection of melanoma by monitoring the intracellular fluorescein fluorescence polarization changes in lymphocytes

The effectiveness of detecting melanoma by measuring the intracellular fluorescein fluorescent polarization (IFFP) of patients' SCM (structuredness of the cytoplasmic matrix)-responding lymphocytes was examined. SCM-responding lymphocytes from 46 melanoma patients and 32 healthy volunteers were labeled with fluorescein diacetate and challenged with different stimuli, and the resulting polarization was determined. The polarizations (P) obtained upon stimulation with nothing (P-0), encephalitogenic factor (P-EF), phytohaemagglutinin (P-PHA), or melanoma antigen (P-MEL), and the ratios RR(ef) (P-EF/P-PHA) and RR(mel) (P-MEL/P-PHA) were lower for SCM-responding lymphocytes from the patients as a group than for those of the controls. The specificity and sensitivity of the IFFP tests (using cutoff values) to detect melanoma were 90.6 and 73.9%, respectively. The IFFP tests may facilitate the discrimination between melanoma patients and healthy subjects, and may be used in follow-up of patients with melanoma.     

Comparison of stochastic approaches to the transportation problem:a case study

The objective of this case study is to provide insight to practitioners about the methodology of using the space-time autoregressive integrated moving average (STARIMA) class of models to formulate stochastic demand of the transportation problem. While providing insight, two other methods-expected value (EV) and stochastic approximation (SA)-are also employed to formulate demand. A comparative evaluation of the methods using brewery data for the distribution of four products from five production plants to 64 distribution centers is presented. It is shown that the demand characterized by the STARIMA approach results in a lower total cost of transportation. This occurs because the STARIMA approach results in better forecasts. Based upon the case study, the cost analysis indicated that the STARIMA method when used without (with) updating resulted in a 9.49% (10.5%) increase in the Company's net profit as compared with the SA method. Similarly, the STARIMA approach when used without (with) updating resulted in an 11.36% (12.37%) increase in the net profit as compared with the EV method. For the STARIMA approach, computations for a large size problem are shown to be identical to those of the deterministic transportation problem given the demand forecasts. Extra computation effort for producing STARIMA forecasts are easily justified in terms of the increased profit margin     

Failure Analysis Approach to Fracture Studies in Powder Metallurgy Parts

This paper presents a failure analysis of tool steel and brass powder metallurgy (P/M) parts that failed during service. A detailed failure investigation of fractured tool steel and brass parts was carried out to assess the causes for their premature failures. The fractured surfaces of the broken pieces and the component surfaces were subjected to detailed examination. Investigations were carried out by visual methods, microhardness measurements, and using optical and scanning electron microscopes. In the case of the brass sample, visual examination of the surface indicated flat surface features. Detailed optical and electron microscopic studies corroborated by microhardness indentations have conclusively established that the failure was mainly due to the presence of very small impurities in the brass component material. In the case of the tool steel sample, the fractured surfaces of the component were subjected to destructive and nondestructive tests. Representative fractured pieces were examined visually and tested for their yield strength using simple tensile tests. Optical and scanning electron microscopies at appropriate magnifications were also performed to characterize its microstructure and fracture morphology. Detailed investigations of the tool steel part established that the failure was mainly due to inferior yield strength of the component resulting from improper heat treatment.     

Modelling and experimental investigation of devolatilizing wood in a fluidized bed combustor

A two-dimensional model is developed for the determination of devolatilization time and char yield of cylindrical wood particles in a bubbling fluidized bed combustor. By using the concept of shape factor, the model is extended to particles of cuboid shape. The model prediction of the devolatilization time agrees with the measured data (present and those reported in the literature) for cylindrical and cuboidal shaped particles within ±20% while the char yield is predicted within ±17%. Influence of some important parameters namely, thermal diffusivity, external heat transfer coefficient and shrinkage, on the devolatilization time and char yield are studied. Thermal diffusivity shows noticeable influence on devolatilization time. The external heat transfer coefficient shows little influence beyond a value of 300 W/(m² K). However particle shrinkage shows negligible effect on the devolatilization time but has a significant influence on the char yield.     

Grafting of methyl methacrylate to nitrocellulose by ceric ions

Studies were carried out on grafting of various vinyl monomers to nitrocellulose by ceric ions. It was observed that graft copolymerization occurred only with methyl methacrylate (MMA) and methyl acrylate monomer. The variables such as initiator concentration, monomer concentration, time of grafting, and nitrocellulose content on grafting of MMA are discussed. By hydrolyzing away the nitrocellulose backbone, the grafted poly(methyl methacrylate) branches were isolated and the >c?o peak at 1740 cm^?1 in the infrared spectra of these isolated branches gave definite evidence of grafting. The molecular weight of isolated branches has been determined by viscometry. The probable mechanism of grafting may be at the α-carbon atom of primary alcohol or at a C₂-C₃ glycol group of the anhydro glucose unit or at the hemiacetal group of the end unit of nitrocellulose, as nitrocellulose is formed by the partial nitration of cotton cellulose.     

Machining characteristics and fracture morphologies in a copper-beryllium (Cu-2Be) alloy

The technology of materials removal is improved greatly by the introduction of advanced cutting tools like cubic boron nitride, ceramics, polycrystalline diamond and the more recent whisker-reinforced materials. In this paper, the influence of cutting temperature on machinability, mechanical properties, microstructure, and fracture morphology of Cu-2Be alloy using a polycrystalline diamond cutter is investigated. The information on machining, microstructure, and fracture morphology of Cu-2Be alloy are very useful to understand their fabrication characteristics and the basic mechanisms of its deformation and fracture. The machinability (in terms of surface finish) of Cu-2Be alloy is evaluated as a function of cutting temperature, resulting from wet and dry cutting. Machining is carried out on a Hardinge Cobra 42 CNC machine (Hardinge Inc., Elmira, NY), and the machining parameters used—cutting speed, depth of cut, and feed rate—are kept constant during both wet and dry cutting. The machined surface finish on Cu-2Be alloy is measured using a surface finish analyzer (Surftest 401, series 178) technique. The machined specimens are examined for their strength and hardness properties using a standard Universal Testing Machine and Rockwell hardness tester, respectively. Wet cutting (using coolants) produced a smooth surface finish when compared with dry cutting of the Cu-2Be alloy. The machined specimens are examined for their microstructural features using a Nikon optical microscope. The specimens are etched using a suitable etchant solution for revealing such microstructure constituents as grain size, phase proportions, and the possible overheated areas (especially in dry cutting). The fractured surfaces from the tensile and impact toughness tests are investigated for their fracture morphologies (dry and wet cutting) using a microprocessor-controlled scanning electron microscope (Jeol Model JSM 5910 LV). A detailed analysis is also made to understand and interpret the basic fracture mechanisms responsible for crack initiation and crack propagation. The Cu-2Be alloy showed relatively higher mechanical properties in wet cutting in comparison to dry cutting operations. Fracture studies demonstrated intergranular and ductile fractures as dominant modes of fracture mechanisms in Cu-2Be alloy.