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A combination of whole cell transport assays and photoaffinity labeling of Pdr5p were used with [(super 125)I]iodoarylazidoprazosoin in purified plasma membrane vesicles to study the behavior of two series of novel substrates namely trityl (triphenylmethyl) and carbazole derivatives. The result showed that site 2, defined initially by tritymidazole efflux requires at least a single hydrogen bond acceptor group.

Research has been conducted on P-glycoprotein, human gene product. The role of conserved glutamate residues in the Walker B domain of the ATP sites has been investigated during human P-glycoprotein catalytic cycle and the details are reported.

the accessibility of the FLAG epitope tag, introduced into the three membrane-spanning domains of the human multidrug resistance protein transporter, MRP1, depends on the location of the tag and the glycosylation sites in the membrane-spanning domains 1 and 3. The cell surface glycosylation may play a role in masking the extracellular epitopes of the transporter.

Research describes the construction of a combinatorial library of polyenes and their use in the discovery of multidrug resistance reversal polyenes. Results indicate that compounds 4ek and 4ej are the most potent polyenes whose interaction with P-glycoprotein involves both ends of the polyenes.

The recombinant baculovirus system was used to express human P-glycoprotein (Pgp), a protein responsible for multidrug resistance because it functions as a drug efflux pump. The activity of purified Pgp from this system is comparable to that of the native enzyme. Analysis of nucleotide binding and hydrolysis using native substrates and various analogues reveal that the binding affinity of Pgp to its substrate is markedly reduced during the catalytic transition state of the enzyme.

A study was conducted on HIV-1 protease inhibitors as substrates for the MDR1 multidrug transporter. The findings indicate that cells in patients who express the MDR1 transporter will be relatively resistant to the anti-viral effects of the HIV-1 protease inhibitors. Moreover, the multidrug transporter may affect the absorption, excretion and distribution of these inhibitors in the body.

The glutathione S-transferase gene fusion system was used to obtain the fusion proteins of the linker region of the human MDR1 gene product. Phosphorylation and phosphopeptide mapping were then conducted to identify specific sites of phosphorylation, which is believed to modulate multidrug resistance by P-glycoprotein. Several serine residues in the linker region were identified as phosphorylation sites for protein kinase C and for P-glycoprotein.

Studies on the human red cell channel-forming integral protein's (CHIP) ultrastructure, pharmacology and transport reveal that CHIP proteoliposomes resemble native red cell water channels. Reconstituted CHIP possess bidirectional orientation and form intramembrane particles. The 24-kDa cleaved CHIP form behaves like the native molecule. CHIP mutants lacking 4-kDa C-terminal domain are inactive in Xenopus laevis oocytes.

A study was conducted on water channel reconsitution in purified red cell CHIP28 protein-containing liposomes. CHIP28 was purified from human red blood cells and reconstituted into proteoliposomes to characterize its physiological behavior and clarify its supposed role as the functional unit of water channels. Results verify that CHIP28 protein does function as a molecular water channel and show that this liposome influences a majority of the transmembrane water movement in red blood cells.

A genetic screen to isolate second-site mutations of S558Y that restore drug transport was used to characterize the transmission interface in the yeast multidrug transporter Pdr5 that uncouples ATP hydrolysis and drug transport. The results offer insight into the deviant and canonical Q-loop residues which are functionally overlapping and equivalent in a strikingly asymmetric ABC transporter.

The occluded nucleotide conformation in wild-type P-glycoprotein (Pgp) which is generated by using nonhydrolyzable ATP analogue ATP-[gamma]-S is described. These studies could be useful to generate pre-hydrolysis [E.S] intermediate in other ATP-binding cassette (ABC) transporters.

The ATPase activity of the yeast Pdr5p transporter protein by its transport substrate clotrimazole is studied. The results demonstrate that Pdr5p can use GTP, as well as ATP to provide energy for substrate transport, hence increasing its multidrug transport substrate specificity.

The investigation of two distinct molecular weight forms of human ABCB6 was observed to be localized to both the outer mitochondrial membrane and the plasma membrane and plays a functional role in the plasma membrane.

Photoaffinity labeling and accumulation assays were used to screen potential substrates or modulators of the human ATP-binding cassette transporter (ABCG2). It was shown that the photoaffinity analogues [125.sup.I] iodoarylazidoprazosin (IAAP) and [3.sup.H]azidopine can be used to photolable both R482-ABCG2 (wild-type) and T482-ABCG2 (mutant).

A study was conducted to assess the role of the conserved Y401 and Y1044 residues of human P-glycoprotein (Pgp) and these residues were substituted F, W, C, or A either singly or together. Systematic biochemical characterization of these residues demonstrated that only aromatic residues at this position allow full retention of function, called the 'A-loop' of the ABC domain, whereas residues such as L or C support only partial (30-50%) activity.

The dimerization motif GXXXG found in the first transmembrane (TM) of the ABCG2 protein is studied by substituting glycine residues with leucine or alanine. The GXXXG motif facilitates dimerization by allowing packing of the helices though it does not induce dimerization single-handedly.

Human P-glycoprotein (Pgp) is an ATP-dependent efflux pump for a variety of chemotherapeutic drugs. The study is conducted to evaluate whether Pgp modulators can be engineered to exhibit high-affinity binding using polyvalency.

A study was conducted to determine a number of residues in transmembrane 12 of P-glycoprotein (Pgp) which are not conserved among known Pgp family members. The vanadate-sensitive release of inorganic phosphate from ATP was determined to measure Pgp-associated ATPase activity. Results indicated that putative amino-proximal half-based nonconserved amino acid residues support a critical role in substrate specificity and drug transport.

Research was conducted to examine the functional significance of the region that links the amino- and carboxyl-terminal halves of P-glycoprotein. The impact of altering the composition and the length of the linker region were investigated by insertion and deletion analysis using a transient vaccinia virus expression system and stable drug selections. Results indicate that a flexible connector region is sufficient for coordinating the function of the two domains.

Wereport on experiments of droplet evaporation on a structuredsuperhydrophobic surface that displays very high contact angle (CA∼ 160 deg), and negligible contact angle hysteresis (<1deg). The droplet evaporation is observed to occur in a constant-contact-anglemode, with contact radius shrinking for almost the entire durationof evaporation. Experiments conducted on Teflon-coated smooth surface(CA ∼ 120 deg) as a baseline also support an evaporation processthat is dominated by a constant-contact-angle mode. The experimentalresults are compared with an isothermal diffusion model for dropletevaporation from the literature. Good agreement is observed for theTeflon-coated smooth surface between the analytical expression andexperimental results in terms of the total time for evaporation, transientvolume, contact angle, and contact radius. However, for the structuredsuperhydrophobic surface, the experiments indicate that the time takenfor complete evaporation of the droplet is greater than the predictedtime, across all droplet volumes. This disparity is attributed primarilyto the evaporative cooling at the droplet interface due to the highaspect ratio of the droplet and also the lower effective thermal conductivityof the substrate due to the presence of air gaps. This hypothesisis verified by numerically evaluating the temperature distributionalong the droplet interface. We propose a generalized relation forpredicting the instantaneous volume of droplets with initial CA >90 deg, irrespective of the mode of evaporation. [ABSTRACT FROM AUTHOR]

We develop and demonstrate the use of electrowetting to achieve the dewetting (Wenzel-to-Cassie transition) of superhydrophobic surfaces. We effect this transition by means of an opposing flat plate and a three-electrode system; the liquid droplet is completely pulled out of its wetted Wenzel state upon the application of a suitable voltage. We also experimentally quantify the dissipative forces preventing the dewetting transition. The energy associated with these nonconservative forces is comparable to the interfacial energies. [ABSTRACT FROM AUTHOR]

Control and switching of liquid droplet states on artificially structured surfaces have significant applications in the field of microfluidics. The present work introduces the concept of using structured surfaces consisting of noncommunicating roughness elements to prevent the transition of a droplet from the Cassie to the Wenzel state. The use of noncommunicating roughness elements leads to a confinement of the medium under the droplet in its Cassie state. Transition to the Wenzel state on such surfaces requires expulsion of this confined medium, which offers significantly increased resistance to the Wenzel transition unlike surfaces consisting of communicating roughness elements. This enhances the robustness of the Cassie state and significantly minimizes the possibility of the Cassie−Wenzel transition. In the present work, the resistance of a surface to the Wenzel transition is measured in terms of the electrowetting (EW) voltage required to trigger this transition. It is seen that surfaces with noncommunicating roughness elements (cratered surfaces) require significantly higher voltages to trigger the Wenzel transition than corresponding surfaces with communicating roughness elements. The findings from the present work also indicate that EW-induced droplet morphology control characteristics show a strong dependence on the nature of the roughness elements (communicating versus noncommunicating). Different aspects of droplet morphology and EW-induced state transition control on surfaces with noncommunicating roughness elements are analyzed; it is seen that such surfaces offer significant possibilities for the development of robust superhydrophobic surfaces. [ABSTRACT FROM AUTHOR]

Electrowetting (EW) has recently been demonstrated as a powerful tool for controlling droplet morphology on smooth and artificially structured surfaces. The present work involves a systematic experimental investigation of the influence of electrowetting in determining and altering the state of a static droplet resting on an artificially microstructured surface. Extensive experimentation is carried out to benchmark a previously developed energy-minimization-based model that analyzed the influence of interfacial energies, surface roughness parameters, and electric fields in determining the apparent contact angle of a droplet in the Cassie and Wenzel states under the influence of an EW voltage. The EW voltage required to trigger a transition from the Cassie state to the Wenzel state is experimentally determined for surfaces having a wide range of surface parameters (surface roughness and fraction of surface area covered with pillars). The reversibility of the Cassie−Wenzel transition upon the removal of the EW voltage is also quantified and analyzed. The experimental results from the present work form the basis for the design of surfaces that enable dynamic control of droplet morphology. A significant finding from the present work is that nonconservative dissipative forces have a significant influence in opposing fluid flow inside the microstructured surface that inhibits reversibility of the Cassie−Wenzel transition. The artificially structured surfaces considered in this work have microscale roughness feature sizes that permits direct visual observation of EW-induced Cassie−Wenzel droplet transition; this is the first reported visual confirmation of EW-induced droplet state transition. [ABSTRACT FROM AUTHOR]

Electrowetting (EW) is a powerful tool to control fluid motion at the microscale and has promising applications in the field of microfluidics. The present work analyzes the influence of an electrowetting voltage in determining and altering the state of a static droplet resting on a rough surface. An energy-minimization-based modeling approach is used to analyze the influence of interfacial energies, surface roughness parameters, and electric fields in determining the apparent contact angle of a droplet in the Cassie and Wenzel states under the influence of an EW voltage. The energy-minimization-based approach is also used to analyze the Cassie−Wenzel transition under the influence of an EW voltage and estimate the energy barrier to transition. The results obtained show that EW is a powerful tool to alter the relative stabilities of the Cassie and Wenzel states and enable dynamic control of droplet morphology on rough surfaces. The versatility and generalized nature of the present modeling approach is highlighted by application to the prediction of the contact angle of a droplet on an electrowetted rough surface consisting of a dielectric layer of nonuniform thickness. [ABSTRACT FROM AUTHOR]