Energetics of assembling an artificial heterodimer with an alpha/beta motif: cleaved versus uncleaved Escherichia coli thioredoxin.

We have studied the folding/binding process between the N- and C-fragments (1-73, 74-108) of oxidized Escherichia coli thioredoxin (Trx) to compare the energetics between the cleaved and uncleaved Trx. Sedimentation equilibrium analysis in 0.1 M potassium phosphate, pH 5.

4-bromotiglic acid, a novel inhibitor of thiolases and a tool for assessing the cooperation between the membrane-bound and soluble beta-oxidation systems of rat liver mitochondria.

An inhibitor of long-chain 3-ketoacyl-CoA thiolase has been developed as a tool for probing the cooperation between the two fatty acid beta-oxidation systems located in the inner mitochondrial membrane and in the mitochondrial matrix, respectively. 4-Bromotiglic acid was synthesized and found to inhibit palmitoylcarnitine-supported respiration of rat liver mitochondria in concentration-dependent and time-dependent fashions. Complete inhibition of respiration was achieved after incubating coupled mitochondria with 10 microM 4-bromotiglic acid for 2 min.

Raman difference studies of GDP and GTP binding to c-Harvey ras.

The vibrational spectra of phosphate modes for GDP and GTP bound to the c-Harvey p21(ras) protein have been determined using 18O isotope edited Raman difference spectroscopy. A number of the phosphate stretch frequencies are changed upon GDP/GTP binding to ras, and the results are analyzed by ab initio calculations and through the use of empirical relationships that relate bond orders and bond lengths to vibrational frequencies. Bound GDP is found to be strongly stabilized by its interactions, mostly electrostatic, with the active site Mg2+.

Raman difference spectroscopic studies of the myosin S1.MgADP.vanadate complex.

The Raman spectra of the nonbridging V--O bonds in the myosin S1.MgADP.Vi complex, often believed to be a transition-state analogue for the phosphotransfer reaction catalyzed by myosin, and in a vanadate solution model compound have been obtained using Raman difference spectroscopic techniques.

Proline isomerization-independent accumulation of an early intermediate and heterogeneity of the folding pathways of a mixed alpha/beta protein, Escherichia coli thioredoxin.

Oxidized Escherichia coli thioredoxin (Trx) is a small protein of 108 residues with one disulfide bond (C32-C35 essentially involved in the activity) and no prosthetic moieties, which folds into a structural motif containing a central twisted beta-sheet flanked by helices that is found in many larger proteins. The kinetics of refolding of Trx in vitro have been investigated using a newly developed active site titration assay and continuous or stopped-flow (SF) methods in conjunction with circular dichroism (CD) and fluorescence (Fl) spectroscopy. These studies revealed the presence of early folding intermediates with "molten globule or pre-molten globule" characteristics.

Stokes problems for moving half-planes.

New exact solutions of the Navier-Stokes equations are obtained for the unbounded and bounded oscillatory and impulsive tangential edgewise motion of touching half-infinite plates in their own plane. In contrast to Stokes classical solutions for the harmonic and impulsive motion of an infinite plane wall, where the solutions are separable or have a simple similarity form, the present solutions have a two-dimensional structure in the near region of the contact between the half-infinite plates. Nevertheless, it is possible to obtain relatively simple closed-form solutions for the flow field in each case by defining new variables which greatly simplify the r- and theta- dependence of the solutions in the vicinity of the contact region.

Raman spectroscopic studies of NAD coenzymes bound to malate dehydrogenases by difference techniques.

We report here on the Raman spectra of NADH, 3-acetylpyridine adenine dinucleotide, APAD+, and a fragment of these molecules, adenosine 5'-diphosphate ribose (ADPR) bound to the mitochondrial (mMDH) and cytoplasmic (or soluble, sMDH) forms of malate dehydrogenase. We observe changes in the Raman spectrum of the adenosine moiety of these cofactors upon binding to mMDH, indicating that the binding site is hydrophobic. On the other hand, there is little change in the spectrum of the adenosine moiety when it binds to sMDH.