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The characterization of active and non-active histidines demonstrates its role in probing the electrostatic interactions along the reaction pathway of a glycoside hydrolase. The results show that histidine play very significant structural and functional roles in the hydrolase because of its diverse ionization behaviors.

A key part in the cytokinetic mechanism of Schizosaccharomyces pombe is the contractile ring myosin, which consists of two heavy chains (Myo2p), two essential light chains (Cdc4p) and two regulatory light chains. NMR spectroscopy is used to compare the structure, stability, and dynamics of the isolated N- and C-terminal domains with one another and with native Cdc4p.

Research has been conducted on endo-beta-1,4-xylanase 10A from Streptomyces lividans xylanase which includes a 130-residue C-terminal family 13 carbohydrate-binding module. The effect of this mulivalency on carbohydrate binding has been investigated via the use of NMR spectroscopy and the results are reported.

Research has been conducted on the endo-beta-(1,4)-xylanase from Bacillus circulans (BCX). Some BCX variants containing active site residue substitutions have been analyzed via NMR spectroscopy and X-ray crystallography and the results are described.

Research has been conducted on the Cellulomonas fimi 1,4-beta-glucanase CenC cellulose-binding domains. The tertiary structure of these domains has been investigated via heteronuclear magnetic resonance spectroscopy.

The resolution structure of the glycosyl-enzyme intermediate is formed on the retaining beta-1,4-xylanase from Bacillus circulans. An analysis of the 1.8 Angstroms resolution structure of the intermediate shows that atoms C5, O5, C1 and C2 of the sugar attains coplanarity as required at the oxocarbenium ion-like transition phases of the double-displacement catalytic mechanism. Several differences beyond the distortion of the sugar moiety are observed following X-ray crystallographic analysis.

A study was conducted to examine the interactions of the N-terminal cellulose-binding domain from Cellulomonas fimi beta-1,4-glucanase CenC with calcium by calorimetry and nuclear magnetic resonance (NMR) spectroscopy. A 50 mM sodium chloride was utilized as a buffer for the NMR studies while a line shape analysis was carried out to determine calcium kinetics. Results indicated that metal ion binding was thermodynamically correlated with native-state stability.

A study was conducted to analyze the thermal stability and oligosaccharide-binding thermodynamics of the N-terminal cellulose-binding domain of the Cellulomonas fimi endoglucanase cenC. Differential scanning calorimetry was utilized to measure stability and oligosaccharide-binding thermodynamics. Results revealed that the intramolecular link in the Cellulose fimi gene contributed to the stability of the endoglucanase cenC and a global two-dimensional minimization routine was utilized to regress the unfolding thermodynamics for the endoglucanase cenC-cellopentaose system.

A research was conducted to study the dynamic behavior of His149 in Bacillus circulans xylanase (BCX) and its relationship with a bound water molecule. Nuclear magnetic resonance spectroscopy was utilized to determine the structure and dynamic environment of the histidine residue. Hydrogen exchange measurements were carried out on unlabeled BCX under varying conditions. Results showed that His149 plays a major role in BCX because of its evolutionary conservation.

The implications for phosphorylation and molecular recognition of Bacillus subtilis response regulator (Spo0F) was investigated by employing high-resolution magnetic resonance structure. Results revealed the presence of relative charge distribution residues located at the site of phosphorylation. This is essential in providing specificity for protein-protein interactions and determining lifetimes of the phosphorylated state.

The structure of the N-terminal cellulose-binding domain from Cellulomonas fimi 1,4-beta-glucanase CenC was determined by multidimensional heteronuclear nuclear magnetic resonance spectroscopy. The results revealed 10 beta-strands which were folded into two antiparallel beta-sheets, forming a jelly-roll beta-sandwich. The strands of the protein surface were shorter than those found in the opposite side of the protein, resulting in a five-stranded binding cleft containing hydrophobic residues at the center and flanked by polar hydrogen-bonding groups.

The N-terminal cellulose-binding domain (CBDN1) from Cellulomonas fimi beta-1,4-glucanase CenC binds amorphous but not crystalline cellulose. The structural and thermodynamic bases of cellulose binding were studied through NMR and difference ultraviolet absorbance spectroscopy in conjunction with calorimetry. The CBDN1 cellulose-binding site spans about five glucosyl units. The cellooligosaccharides bind along a five-stranded beta-sheet that forms a concave face of the jelly-roll beta-sandwich structure of CBDN1.

Selective isotopic labelling and 13C-NMR studies on the pK(sub a) values of the catalytic glutamic acid residues in wild type/mutant Bacillus circulans (BCX) and subtilis xylanases reveal a two-step hydrolysis mechanism involving a covalent glycosyl-enzyme intermediate. The double-displacement reaction of BCX may be broken into glycosylation and deglycosylation steps. The glycosylation step produces the glycosyl-enzyme intermediate and deglycosylation regenerates the native enzyme. These protonation and deprotonation steps are governed by glutamic ionization states.

Analysis of the secondary structure of the ETS domain of murine Ets-1 by FT-IR spectroscopy, NMR chemical shifts and circular dichroism technique reveals that the ETS domain contains four beta-strands and three alpha-helices and that the ets proteins belong to the winged helix-turn-helix DNA-binding protein family. The four-stranded antiparallel beta-sheet functions as the scaffold for the putative helix-turn-helix DNA recognition motif. The Ets proteins have the same topology as heat shock factor, catabolite activator protein and hepatocyte nuclear factor.

X-ray crystallography and NMR spectroscopy were used to determine and probe the structure of circular permutant of Bacillus circulans xylanase. The results for the circularly permuted xylanases provides an excellent set of new start points for directed evolution of commercially important Bcx enzyme and also provides new insigth into the mechanism of enzymatic glycoside hydrolysis.

NMR spectroscopy was used to characterize the crystal structure and the dynamics of the GT family 42 Campylobacter jejuni [alpha]-2,3/2,8-sialyltransferase (CstII) and measure the intrinsic p[K.sub.a] value of His188 for comparison with the p[K.sub.a] determined from the pH-dependent [k.sub.cat]/[K.sub.M] of the enzyme. The site-specific p[K.sub.a] value ~6.6 of His188 was observed to match the apparent p[K.sub.a] value ~6.5 which governed the pH dependence of [k.sub.cat]/[K.sub.M] for CstII toward CMP-Neu5Ac in the presence of saturating acceptor substrate.

The studies have shown that [Ca.sup.2+]-ligated calmodulin (CaM) encompassing EF-hands 2 and 3 (CaM2/3) are bound to a peptide representing the CaM-recognition sequence of skeletal muscle myosin light chain kinase (M13). They have highlighted the plasticity of EF-hand association and the diverse repertoire of mechanisms possible for CaM-target protein interactions.

[(super 1)H] and [(super 15)H] correlation spectroscopy is used to examine the lysine amines in CexCD and its trapped glycosyl-enzyme intermediate, demonstrating clearly that K47 interacts with the cellobiosyl moiety via its positively charged side chain. It is shown that in cursory survey, well-resolved amine signals were observed in the [(super 1)H]- [(super 15)H] HMQC spectra of ubiquitin, RanGAP, and a xylanase.