Background: Low-cost handheld computers (PDA) potentially represent an efficient tool for collecting sensitive data in surveys. The goal of this study is to evaluate the quality of sexual behavior data collected with handheld computers in comparison with paper-based questionnaires.

This paper describes and illustrates the use of ensemble-based docking, i.e., using a collection of protein structures in docking calculations for hit discovery, the exploration of biochemical pathways and toxicity prediction of drug candidates. We describe the computational engineering work necessary to enable large ensemble docking campaigns on supercomputers. We show examples where ensemble-based docking has significantly increased the number and the diversity of validated drug candidates. Finally, we illustrate how ensemble-based docking can be extended beyond hit discovery and toward providing a structural basis for the prediction of metabolism and off-target binding relevant to pre-clinical and clinical trials.

Abstract. Pyrazinamidase of Mycobacterium tuberculosis catalyzes the conversion of pyrazinamide to the active molecule pyrazinoic acid. Reduction of pyrazinamidase activity results in a level of pyrazinamide resistance. Previous studies have suggested that pyrazinamidase has a metal-binding site and that a divalent metal cofactor is required for activity. To determine the effect of divalent metals on the pyrazinamidase, the recombinant wild-type pyrazinamidase corresponding to the H37Rv pyrazinamide-susceptible reference strain was expressed in Escherichia coli with and without a carboxy terminal. His-tagged pyrazinamidase was inactivated by metal depletion and reactivated by titration with divalent metals. , and Mg 2+ did not restore the activity under the conditions tested. Various recombinant mutated pyrazinamidases with appropriate folding but different enzymatic activities showed a differential pattern of recovered activity. X-ray fluorescence and atomic absorbance spectroscopy showed that recombinant wild-type pyrazinamidase expressed in E. coli most likely contained Zn. In conclusion, this study suggests that M. tuberculosis pyrazinamidase is a metalloenzyme that is able to coordinate several ions, but in vivo, it is more likely to coordinate Zn 2+ . However, in vitro, the metal-depleted enzyme could be reactivated by several divalent metals with higher efficiency than Zn.

The Escherichia coli cAMP receptor protein, CRP, is a homodimeric global transcription activator that employs multiple mechanisms to modulate the expression of hundreds of genes. These mechanisms require different interfacial interactions among CRP, RNA, and DNA of varying sequences. The involvement of such a multiplicity of interfaces requires a tight control to ensure the desired phenotype. CRP-dependent promoters can be grouped into three classes. For decades scientists in the field have been puzzled over the differences in mechanisms between class I and II promoters. Using a new crystal structure, IR spectroscopy, and computational analysis, we defined the energy landscapes of WT and 14 mutated CRPs to determine how a homodimeric protein can distinguish nonpalindromic DNA sequences and facilitate communication between residues located in three different activation regions (AR) in CRP that are ϳ30 Å apart. We showed that each mutation imparts differential effects on stability among the subunits and domains in CRP. Consequently, the energetic landscapes of subunits and domains are different, and CRP is asymmetric. Hence, the same mutation can exert different effects on ARs in class I or II promoters. The effect of a mutation is transmitted through a network by long-distance communication not necessarily relying on physical contacts between adjacent residues. The mechanism is simply the sum of the consequences of modulating the synchrony of dynamic motions of residues at a distance, leading to differential effects on ARs in different subunits. The computational analysis is applicable to any system and potentially with predictive capability. The Escherichia coli cAMP receptor protein (CRP), 5 is a homodimeric global transcription activator that controls the transcription of numerous genes involving carbon utilization (1, 2). The CRP-dependent promoters can be grouped into three classes (3-5). In class I promoters, CRP binds upstream of RNA polymerase (RNAP), at a site centered close to position Ϫ61.5 on the DNA (3, 6). In class II promoters, CRP is sandwiched between the ␣-CTD and 70 subunit of RNAP, at a site centered close to position Ϫ41.5 on the DNA (4, 7). In class III promoters, two or more CRP dimers bind at multiple sites through class I or a combination of class I/class II CRP-dependent promoters (5, 6). Fig. 1 shows the models of class I and II promoters with respect to the relative positions of CRP, RNAP, and the two DNA half-sites for CRP binding. Three surfaceexposed regions of CRP, designated as activating regions 1-3 (AR 1 , AR 2 , and AR 3), modulate the contacts with RNAP (4). The contacts between ARs and RNAP are promoter-dependent. Fig. 2A shows the relative locations of these surfaces in CRP and the distances between key residues in these surfaces. The architectural designs of these promoters and the extensive results of mutation studies of class II promoters (4-6, 8-13) enabled the field to conclude that RNAP contacts AR 1 of the CRP upstream subunit. Furthermore, through analogous modeling, ...