The transfer of electrons between proteins is an essential step in biological energy production. Two protein redox partners are often artificially crosslinked to investigate the poorly understood... Show moreThe transfer of electrons between proteins is an essential step in biological energy production. Two protein redox partners are often artificially crosslinked to investigate the poorly understood mechanism by which they interact. To better understand the effect of crosslinking on electron transfer rates, we have constructed dimers of azurin by crosslinking the monomers. The measured electron exchange rates, combined with crystal structures of the dimers, demonstrate that the length of the linker can have a dramatic effect on the structure of the dimer and the electron transfer rate. The presence of ordered water molecules in the protein protein interface may considerably influence the electronic coupling between redox centers. Show less
L-type Ca(2+) channels are an important means by which a cell regulates the Ca(2+) influx into the cytosol on electrical stimulation. Their structure and dynamics in the plasma membrane, including... Show moreL-type Ca(2+) channels are an important means by which a cell regulates the Ca(2+) influx into the cytosol on electrical stimulation. Their structure and dynamics in the plasma membrane, including their molecular mobility and aggregation, is of key interest for the in-depth understanding of their function. Construction of a fluorescent variant by fusion of the yellow-fluorescent protein to the ion channel and expression in a human cell line allowed us to address its dynamic embedding in the membrane at the level of individual channels in vivo. We report on the observation of individual fluorescence-labeled human cardiac L-type Ca(2+) channels using wide-field fluorescence microscopy in living cells. Our fluorescence and electrophysiological data indicate that L-type Ca(2+) channels tend to form larger aggregates which are mobile in the plasma membrane. Show less
There is no agreement, in the chick, about the number of the endocardial cushions within the outflow tract or their pattern of fusion. Also, little is known of their relative contributions to the... Show moreThere is no agreement, in the chick, about the number of the endocardial cushions within the outflow tract or their pattern of fusion. Also, little is known of their relative contributions to the formation of the arterial valves, the subpulmonary infundibulum, and the arterial valvar sinuses. As the chick heart is an important model for studying septation of the outflow tract, our objective was to clarify these issues. Normal septation of the outflow tract was studied in a series of 60 staged chick hearts, by using stained whole-mount preparations, serial sections, and scanning electron microscopy. A further six hearts were examined subsequent to hatching. At stage 21, two pairs of endocardial cushions were seen within the developing outflow tract. One pair was positioned proximally, with the other pair located distally. By stage 25, a third distal cushion had developed. This finding was before the appearance of two further, intercalated, endocardial cushions, also distally positioned, which were first seen at stage 29. In the arterial segment, the aortic and pulmonary channels were separated by the structure known as the aortopulmonary septum. The dorsal limb of this septum penetrated the distal dorsal cushion, whereas the ventral limb grew between the remaining two distal cushions, both of which were positioned ventrally. The three distal endocardial cushions, and the two intercalated endocardial cushions, contributed to the formation of the leaflets and sinuses of the arterial roots. The two proximal cushions gave rise to a transient septum, which later became transformed into the muscular component of the subpulmonary infundibulum. Concomitant with these changes, an extracardiac tissue plane was formed which separated this newly formed structure from the sinuses of the aortic root. Our study confirms that three endocardial cushions are positioned distally, and two proximally, within the developing outflow tract of the chick. The pattern of the distal cushions, and the position of the ventral limb of the aortopulmonary septum, differs significantly from that seen in mammals. Anat Rec 264:273-283, 2001. (C) 2001 Wiley-Liss, Inc. Show less
The understanding of the interplay of electron correlations and randomness in solids is enhanced by demonstrating that particle-hole ( p−h) symmetry plays a crucial role in determining the effects... Show moreThe understanding of the interplay of electron correlations and randomness in solids is enhanced by demonstrating that particle-hole ( p−h) symmetry plays a crucial role in determining the effects of disorder on the transport and thermodynamic properties of the half-filled Hubbard Hamiltonian. We show that the low-temperature conductivity decreases with increasing disorder when p−h symmetry is preserved, and shows the opposite behavior, i.e., conductivity increases with increasing disorder, when p−h symmetry is broken. The Mott insulating gap is insensitive to weak disorder when there is p−h symmetry, whereas in its absence the gap diminishes with increasing disorder. Show less
Chin-A-Woeng, T.F.C.; Broek, D. van den; Voer, G. de; Drift, K.M.G.M. van der; Tuinman, S.; Thomas-Oates, J.E.; ... ; Bloemberg, G.V. 2001
An optical single-molecule study is reported of a quickly frozen solution of 2.3,8.9-dibenzanthanthrene (DBATT) in n-tetradecane at 1.4 K. The orientation has been measured of several hundreds of... Show moreAn optical single-molecule study is reported of a quickly frozen solution of 2.3,8.9-dibenzanthanthrene (DBATT) in n-tetradecane at 1.4 K. The orientation has been measured of several hundreds of DBATT molecules within a confocal detection volume of similar to 10 mum(3) as a function of their resonance frequency in the range of the two 0-0 bands in the fluorescence-excitation spectrum. Each band is found to correspond to a distinct distribution of orientations of DBATT molecules. A particular resonance frequency within a band is not correlated with a specific molecular orientation. (C) 2001 Elsevier Science B.V. All rights reserved. Show less
Decreased phosphorylation of focal adhesion kinase (FAK) is associated with loss of focal adhesions and actin stress fibers and precedes the onset of apoptosis in renal epithelial cells caused by... Show moreDecreased phosphorylation of focal adhesion kinase (FAK) is associated with loss of focal adhesions and actin stress fibers and precedes the onset of apoptosis in renal epithelial cells caused by nephrotoxicants (Van de Water, B., Nagelkerke, J. F., and Stevens, J. L. (1999) J. Biol. Chem. 274, 13328-13337). The role of FAK in the control of apoptosis caused by nephrotoxicants was further investigated in LLC-PK1 cells that were stably transfected with either green fluorescent protein (GFP)-FAK or dominant negative acting deletion mutants of FAK, GFP-FAT, and GFP-FRNK. GFP-FAT and GFP-FRNK delayed the formation of focal adhesions and prevented the localization of endogenous (phosphorylated) FAK at these sites. GFP-FAT and GFP-FRNK overexpression potentiated the onset of apoptosis caused by the nephrotoxicant dichlorovinyl-cysteine. This was associated with an increased activation of caspase-3. GFP-FAT also potentiated apoptosis caused by doxorubicin but not cisplatin. The potentiation of apoptosis by GFP-FAT was related to an almost complete dephosphorylation of FAK; this did not occur in cells overexpressing only GFP. This dephosphorylation was associated with a pronounced loss of focal adhesion organization in GFP-FAT cells, in association with loss of tyrosine phosphorylation of paxillin. In conclusion, the data indicate an important role of cell-matrix signaling in the control of chemically induced apoptosis; loss of FAK activity caused by toxic chemicals results in perturbations of focal adhesion organization with a subsequent inactivation of associated (signaling) molecules and loss of survival signaling. Show less
