Parallel measurements of the thermodynamics (free-energy, enthalpy, entropy and heat-capac change) of ligand binding to FK506 binding protein First suggested >70 years ago (1, 2) and particularly emphasized by Pauling et al. (3,4), the hydrogen bond is now recognized as an interaction of fundamental importance in determining the structures of proteins and their complexes with ligands (5). Nevertheless, conflicting views are still held on the relative energetic contributions ofhydrogen bonds and interactions involving nonpolar groups to the thermodynamics of protein folding and ligand binding (6-8). Models have been presented arguing that the overall enthalpic contribution of interactions involving nonpolar moieties (at 250C) to the process of protein folding is highly unfavorable (7, 9), highly favorable (8), or essentially zero (10, 11). Arguments have also been advanced stating that the overall contribution of amide hydrogen-bond formation to the enthalpy of protein folding (or ligand-protein binding) at 250C is favorable (7), negligible (12) and possibly unfavorable (8,13). Clarification of the present controversy requires estimates of the enthalpies of formation of specific hydrogen bonds in particular regions of known protein structures. Such measurements are essential for understanding the energetic basis of protein folding and for obtaining structure-based predictions of the energies of ligand binding for the purpose of drug design.Part of the difficulty in dissecting the relative energetic contribution of hydrogen bonds to folding or binding reactions of proteins is that the formation of hydrogen bonds between atoms in reaction products is often accompanied by the release ofwater molecules that were hydrogen-bonded to these atoms prior to reaction. Tacrolimus (also known as FK506) and rapamycin are large macrocyclic compounds each of which binds with high affinity to a common cytosolic protein of 12 kDa known as FK506 binding protein . Recently, the structures of FKBP-12 in solution and in the crystalline state have been determined (14, 15), as have the crystal structures of the tacrolimus-FKBP-12 and rapamyci-FKBP-12 complexes (16-18). One of the structural features common to both protein-ligand complexes is a hydrogen bond between the Tyr-82 hydroxyl hydrogen (donor) and the amide carbonyl oxygen (acceptor) of either rapamycin or tacrolimus, buried in the hydrophobic interface (Fig. 1). As determined by x-ray crystallography, an important feature of the unliganded protein structure is the welldefined hydration of this tyrosine hydroxyl group. Furthermore, the NMR structure of tacrolimus in chloroform has been reported by Karuso et al. (19); the low solubility of tacrolimus prevents the determination of its structure in water. However, the interaction of unbound tacrolimus with water has been investigated through molecular dynamics simulation (20). The simulation reveals that the amide carbonyl oxygen of tacrolimus is exposed to solvent and makes an average of 1.6 hydrogen bonds with water mol...

To investigate clinical aspects of the neutropenia induced by high-dose intravenous immunoglobulin therapy (IVIG) we performed serial hematology, including differentiation of white blood cells (WBC), before and after 22 instances of IVIG in 16 patients with neuroimmunologic disorders. WBC and neutrophils showed a significant decrease with a nadir 2 days after IVIG, but returned to previous values by 14 days with no treatment except in 2 cases. No patient showed any infectious complication. Both WBC and neutrophils were significantly decreased in cases without corticosteroid therapy but not in those with medication. In nine instances (4 with and 5 without corticosteroid treatment), CD11b and CD16 on neutrophils were investigated using flow cytometry. In 3 of 5 instances without corticosteroid treatment the expression of CD11b was decreased after IVIG, while no change was detected in CD16. There was no difference in either CD11b or CD16 between before and after IVIG in instances with corticosteroid therapy. Neutropenia commonly and transiently develops just after IVIG, and can be prevented by corticosteroid pretreatment. Circulating neutrophils might bind to the vascular wall mainly with the involvement of CD11b and migrate into a storage pool, resulting in an apparent neutropenia after IVIG.

Aldolases are enzymes that catalyze stereospecific aldol reactions in a reversible manner. Naturally occurring aldolases include class I aldolases, which catalyze aldol reactions via enamine intermediates, and class II aldolases, in which Zn(2+) enolates of substrates react with acceptor aldehydes. In this work, Zn(2+) complexes of L-prolyl-pendant[15]aneN(5) (ZnL(3)), L-prolyl-pendant[12]aneN(4) (ZnL(4)), and L-valyl-pendant[12]aneN(4) (ZnL(5)) were designed and synthesized for use as chiral catalysts for enantioselective aldol reactions. The complexation constants for L(3) to L(5) with Zn(2+) [logK(s)(ZnL)] were determined to be 14.1 (for ZnL(3)), 7.6 (for ZnL(4)), and 9.6 (for ZnL(5)), indicating that ZnL(3) is more stable than ZnL(4) and ZnL(5). The deprotonation constants of Zn(2+)-bound water [pK(a)(ZnL) values] for ZnL(3), ZnL(4), and ZnL(5) were calculated to be 9.2 (for ZnL(3)), 8.2 (for ZnL(4)), and 8.6 (for ZnL(5)), suggesting that the Zn(2+) ions in ZnL(3) is a less acidic Lewis acid than in ZnL(4) and ZnL(5). These values also indicated that the amino groups on the side chains weakly coordinate to Zn(2+). We carried out aldol reactions between acetone and 2-chlorobenzaldehyde and other aldehydes in the presence of catalytic amounts of the chiral Zn(2+) complexes in acetone/H(2)O at 25 and 37 degrees C. Whereas ZnL(3) yielded the aldol product in 43% yield and 1% ee (R), ZnL(4) and ZnL(5) afforded good chemical yields and high enantioselectivities of up to 89% ee (R). UV titrations of proline and ZnL(4) with acetylacetone (acac) in DMSO/H(2)O (1:2) indicate that ZnL(4) facilitates the formation of the ZnL(4)(acac)(-) complex (K(app)=2.1x10(2) M(-1)), whereas L-proline forms a Schiff base with acac with a very small equilibrium constant. These results suggest that the amino acid components and the Zn(2+) ions in ZnL(4) and ZnL(5) function in a cooperative manner to generate the Zn(2+)-enolate of acetone, thus permitting efficient enantioselective C-C bond formation with aldehydes.

An analog hadron calorimeter (AHCAL) prototype of 5.3 nuclear interaction lengths thickness has been constructed by members of the CALICE Collaboration. The AHCAL prototype consists of a 38-layer sandwich structure of steel plates and highly-segmented scintillator tiles that are read out by wavelength-shifting fibers coupled to SiPMs. The signal is amplified and shaped with a custom-designed ASIC. A calibration/monitoring system based on LED light was developed to monitor the SiPM gain and to measure the full SiPM response curve in order to correct for non-linearity. Ultimately, the physics goals are the study of hadron shower shapes and testing the concept of particle flow. The technical goal consists of measuring the performance and reliability of 7608 SiPMs. The AHCAL was commissioned in test beams at DESY and CERN. The entire prototype was completed in 2007 and recorded hadron showers, electron showers and muons at different energies and incident angles in test beams at CERN and Fermilab.

The entire nucleotide sequence was determined for eight hepatitis B virus (HBV) genomes from three symptom-free carriers, two patients with chronic persistent hepatitis and one patient with chronic active hepatitis, who were positive for antibody to hepatitis B e antigen (HBeAg). The two patients with chronic persistent hepatitis were tested again after they developed chronic active or fulminant hepatitis, making a total of eight samples. Six had a point mutation in the preC region prohibiting the encoding of HBeAg precursor, while the remaining two had a deletion of 8 or 21 nucleotides within the X gene upstream of the preC transcription initiation sites which would affect the X gene and the putative preC/C promoter. Most genomes from the three symptom-free carriers and the two patients with chronic persistent hepatitis, with HBV DNA levels of 10(2)-10(3)/ml, had deletion, frameshift mutation, initiation failure or a premature stop codon, rendering them replication-incompetent. In contrast, such mutations were rarely seen in HBV genomes from the two patients with chronic persistent hepatitis after they had developed active or fulminant hepatitis and from the patient with chronic active hepatitis, all of whom had vigorous HBV replication with serum HBV DNA from 10(6) to 10(9)/ml. Unique mutations for amino acid changes were more frequent in HBV genomes with a higher replicative activity. These results indicate two kinds of HBV genomes with an HBeAg-minus phenotype, one with defects seriously affecting viral replication and the other without such defects, which would account for different clinical profiles in carriers with antibody to HBeAg.