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Journal Issue - Volume 17 Issue 8 (August 2008)

Abstract Although phenomenlogical models that account for cooperativity in allosteric systems date back to the early and mid‐60's (e.g., the KNF and MWC models), there is resurgent interest in the topic due to the recent experimental and computational studies that attempted to reveal, at an atomistic level, how allostery actually works. In this review, using systems for which atomistic simulations have been carried out in our groups...

Abstract This review compares the folding behavior of proteins and RNAs. Topics covered include the role of topology in the determination of folding rates, major folding events including collapse, properties of denatured states, pathway heterogeneity, and the influence of the mode of initiation on the folding pathway.

Abstract Protein self‐association is critical to many biological functions. However, atomic‐level structural characterization of these assemblies has remained elusive. In this report we present insights into the mechanistic details of the process of self‐association of the 136‐residue GTPase effector domain (GED) of the endocytic protein dynamin into a megadalton‐sized soluble mass. Our approach is based on NMR monitoring of...

Abstract We describe the use of four complementary biosensors (Biacore 3000, Octet QK, ProteOn XPR36, and KinExA 3000) in characterizing the kinetics of human nerve growth factor (NGF) binding to a humanized NGF‐neutralizing monoclonal antibody (tanezumab, formerly known as RN624). Tanezumab is a clinical candidate as a therapy for chronic pain. Our measurements were consistent with the NGF/tanezumab binding affinity being tighter...

Abstract The genome of Pyrococcus abyssi contains two open reading frames encoding proteins which had been previously predicted to be DNA ligases, Pab2002 and Pab1020. We show that while the former is indeed a DNA ligase, Pab1020 had no effect on the substrate deoxyoligo‐ribonucleotides tested. Instead, Pab1020 catalyzes the nucleotidylation of oligo‐ribonucleotides in an ATP‐dependent reaction, suggesting that it is an RNA ligase....

Abstract In this work, we re‐examine the previously reported phenomenon of the creation of a superactive glutamate dehydrogenase by proteolytic modification by chymotrypsin and explore the various discrepancies that came to light during those studies. We find that superactivation is caused by cleavage at the N terminus of the protein and not the C‐terminal allosteric site, as has previously been suggested. N‐terminal sequencing...

Abstract Although protein Z (PZ) has a domain arrangement similar to the essential coagulation proteins FVII, FIX, FX, and protein C, its serine protease (SP)‐like domain is incomplete and does not exhibit proteolytic activity. We have generated a trial sequence of putative activated protein Z (PZa) by identifying amino acid mutations in the SP‐like domain that might reasonably resurrect the serine protease catalytic activity of PZ....

Abstract Dioxygenases catalyze dioxygen incorporation into various organic compounds and play a key role in the complex degradation pathway of mono‐ and polycyclic aromatic and hetero‐aromatic compounds. Here we report the crystal structure of gentisate 1,2‐dioxygenase from Silicibacter pomeroyi (GDOsp) at a 2.8 Å resolution. The enzyme possessed a conserved three‐dimensional structure of the bicupin family, forming a...

Abstract Newly determined protein structures are classified to belong to a new fold, if the structures are sufficiently dissimilar from all other so far known protein structures. To analyze structural similarities of proteins, structure alignment tools are used. We demonstrate that the usage of nonsequential structure alignment tools, which neglect the polypeptide chain connectivity, can yield structure alignments with significant...

Abstract Cellobiohydrolase from Melanocarpus albomyces (Cel7B) is a thermostable, single‐module, cellulose‐degrading enzyme. It has relatively low catalytic activity under normal temperatures, which allows structural studies of the binding of unmodified substrates to the native enzyme. In this study, we have determined the crystal structure of native Ma Cel7B free and in complex with three different cello‐oligomers: cellobiose (Glc2),...

Abstract We have studied the interaction of the enzyme tissue transglutaminase (tTG), catalyzing cross‐link formation between protein‐bound glutamine residues and primary amines, with Parkinson's disease‐associated α‐synuclein protein variants at physiologically relevant concentrations. We have, for the first time, determined binding affinities of tTG for wild‐type and mutant α‐synucleins using surface plasmon resonance approaches,...

Abstract Human glucose 6‐phosphate dehydrogenase, purified after overexpression in E. coli, was shown to contain one molecule/subunit of acid‐extractable “structural” NADP+ and no NADPH. This tightly bound NADP+ was reduced by G6P, presumably following migration to the catalytic site. Gel‐filtration yielded apoenzyme, devoid of bound NADP+ but, surprisingly, still fully active. Mr of the main component of “stripped” enzyme by gel filtration was ∼100,000,...

Abstract A novel short‐chain (S)‐1‐phenyl‐1,2‐ethanediol dehydrogenase (SCR) from Candida parapsilosis exhibits coenzyme specificity for NADPH over NADH. It catalyzes an anti‐Prelog type reaction to reduce 2‐hydroxyacetophenone into (S)‐1‐phenyl‐1,2‐ethanediol. The coding gene was overexpressed in Escherichia coli and the purified protein was crystallized. The crystal structure of the apo‐form was solved to 2.7 Å resolution. This protein forms...

Abstract The presence of native contacts in the denatured state of many proteins suggests that elements of the biologically active structure of these molecules are formed during the initial stage of the folding process. The rapidity with which these events take place makes it difficult to study them in vitro, but, by the same token, suitable for studies in silico. With the help of all‐atom, explicit solvent, molecular dynamics...

Abstract Amide proton NMR signals from the N‐terminal domain of monomeric α‐synuclein (αS) are lost when the sample temperature is raised from 10°C to 35°C at pH 7.4. Although the temperature‐induced effects have been attributed to conformational exchange caused by an increase in α‐helix structure, we show that the loss of signals is due to fast amide proton exchange. At low ionic strength, hydrogen exchange rates are faster for the...

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