Crystal structure of the MukB hinge domain with coiled‐coil stretches and its functional implications

The structural maintenance of chromosomes (SMC) family proteins are commonly found in the multiprotein complexes involved in chromosome organization, including chromosome condensation and sister chromatid cohesion. These proteins are characterized by forming a V‐shaped homo‐ or heterodimeric structure with two long coiled‐coil arms having two ATPase head domains at the distal ends. The hinge domain, located in the middle of the coiled coil, forms the dimer interface. In addition to being the dimerization module, SMC hinges appear to play other roles, including the gateway function for DNA entry into the cohesin complex. Herein, we report the homodimeric structure of the hinge domain of Escherichia coli MukB, which forms a prokaryotic condensin complex with two non‐SMC subunits, MukE and MukF. In contrast with SMC hinge of Thermotoga maritima which has a sizable central hole at the dimer interface, MukB hinge forms a constricted dimer interface lacking a hole. Under our assay conditions, MukB hinge does not interact with DNA in accordance with the absence of a notable positively charged surface patch. The function of MukB hinge appears to be limited to dimerization of two copies of MukB molecules. Proteins 2010. © 2009 Wiley‐Liss, Inc.     

Canine morphology in the larger Felidae: implications for feeding ecology

Canine morphology is analysed at seven intervals along the crown in both anteroposterior and lateromedial perspective in seven species of large felids. The puma and the snow leopard have stout, rather conical canines, whereas those of lions, jaguars, and tigers bear substantial resemblance to each other, reflecting their phylogenetic relationships, and are less conical and large. The canines of the leopard are intermediate in morphology between those of the other species, probably reflecting its more generalized diet. The clouded leopard has very large and blade-like canines, which are different from the other analysed species. Canine bending strengths to estimated bite forces appear to differs less among the species than morphology, indicating that the evolution of canines has been constricted with respect to their strength in failure, probably owing to their being equally important for species fitness. However, the clouded leopard again stands out, having a high estimated bite force and rather weak canines in bending about the anteroposterior as well as lateromedial planes compared to the other species. Canine morphology to some extent reflects differences in killing mode, but also appears to be related to the phylogeny. The marked divergence of the clouded leopard is presently not understood.  © 2007 The Linnean Society of London, Biological Journal of the Linnean Society , 2007, 91 , 573–592.     

Cholesterol-induced protein sorting: an analysis of energetic feasibility

The mechanism(s) underlying the sorting of integral membrane proteins between the Golgi complex and the plasma membrane remain uncertain because no specific Golgi retention signal has been found. Moreover one can alter a protein's eventual localization simply by altering the length of its transmembrane domain (TMD). M. S. Bretscher and S. Munro (SCIENCE: 261:1280-1281, 1993) therefore proposed a physical sorting mechanism based on the hydrophobic match between the proteins' TMD and the bilayer thickness, in which cholesterol would regulate protein sorting by increasing the lipid bilayer thickness. In this model, Golgi proteins with short TMDs would be excluded from cholesterol-enriched domains (lipid rafts) that are incorporated into transport vesicles destined for the plasma membrane. Although attractive, this model remains unproven. We therefore evaluated the energetic feasibility of a cholesterol-dependent sorting process using the theory of elastic liquid crystal deformations. We show that the distribution of proteins between cholesterol-enriched and cholesterol-poor bilayer domains can be regulated by cholesterol-induced changes in the bilayer physical properties. Changes in bilayer thickness per se, however, have only a modest effect on sorting; the major effect arises because cholesterol changes also the bilayer material properties, which augments the energetic penalty for incorporating short TMDs into cholesterol-enriched domains. We conclude that cholesterol-induced changes in the bilayer physical properties allow for effective and accurate sorting which will be important generally for protein partitioning between different membrane domains.     

A unified model for the origin of DNA sequence-directed curvature

The fine structure of the DNA double helix and a number of its physical properties depend upon nucleotide sequence. This includes minor groove width, the propensity to undergo the B-form to A-form transition, sequence-directed curvature, and cation localization. Despite the multitude of studies conducted on DNA, it is still difficult to appreciate how these fundamental properties are linked to each other at the level of nucleotide sequence. We demonstrate that several sequence-dependent properties of DNA can be attributed, at least in part, to the sequence-specific localization of cations in the major and minor grooves. We also show that effects of cation localization on DNA structure are easier to understand if we divide all DNA sequences into three principal groups: A-tracts, G-tracts, and generic DNA. The A-tract group of sequences has a peculiar helical structure (i.e., B*-form) with an unusually narrow minor groove and high base-pair propeller twist. Both experimental and theoretical studies have provided evidence that the B*-form helical structure of A-tracts requires cations to be localized in the minor groove. G-tracts, on the other hand, have a propensity to undergo the B-form to A-form transition with increasing ionic strength. This property of G-tracts is directly connected to the observation that cations are preferentially localized in the major groove of G-tract sequences. Generic DNA, which represents the vast majority of DNA sequences, has a more balanced occupation of the major and minor grooves by cations than A-tracts or G-tracts and is thereby stabilized in the canonical B-form helix. Thus, DNA secondary structure can be viewed as a tug of war between the major and minor grooves for cations, with A-tracts and G-tracts each having one groove that dominates the other for cation localization. Finally, the sequence-directed curvature caused by A-tracts and G-tracts can, in both cases, be explained by the cation-dependent mismatch of A-tract and G-tract helical structures with the canonical B-form helix of generic DNA (i.e., a cation-dependent junction model).     

Kerr Constant of Vesicle-Like Droplets

The Kerr effect on vesicle-likedroplets (vesicles, cells or emulsion droplets) is described. Wegive a derivation of the Kerr constant for a dielectric fluiddroplet immersed in another fluid, assuming that the droplet in aweak electric field becomes a prolate ellipsoid. The Kerr constantis evaluated also for a droplet covered by a membrane of nonzerothickness. Comparing the theory with the experiment on dropletmicroemulsions from the literature, the bending rigidity constantof the surface layer is estimated.     

Predictions of protein flexibility: first-order measures

The normal modes of a molecule are utilized, in conjunction with classical conformal vector field theory, to define a function that measures the capability of the molecule to deform at each of its residues. An efficient algorithm is presented to calculate the local chain deformability from the set of normal modes of vibration. This is done by considering each mode as an off-grid sample of a deformation vector field. Predictions of deformability are compared with experimental data in the form of dihedral angle differences between two conformations of ten kinases by using a modified correlation function. Deformability calculations correlate well with experimental results and validate the applicability of this method to protein flexibility predictions.     

Bending and flexibility of methylated and unmethylated EcoRI DNA

We used cyclization kinetics experiments and Monte Carlo simulations to determine a structural model for a DNA decamer containing the EcoRI restriction site. Our findings agree well with recent crystal and NMR structures of the EcoRI dodecamer, where an overall bend of seven degrees is distributed symmetrically over the molecule. Monte Carlo simulations indicate that the sequence has a higher flexibility, assumed to be isotropic, compared to that of a "generic" DNA sequence. This model was used as a starting point for the investigation of the effect of cytosine methylation on DNA bending and flexibility. While methylation did not affect bend magnitude or direction, it resulted in a reduction in bending flexibility and under-winding of the methylated nucleotides. We demonstrate that our approach can augment the understanding of DNA structure and dynamics by adding information about the global structure and flexibility of the sequence. We also show that cyclization kinetics can be used to study the properties of modified nucleotides.     

Dynamics of supercoiled and linear pTZ18U plasmids observed with a long-lifetime metal-ligand complex

The metal-ligand complex, [Ru(bpy)2(dppz)]2+ (bpy = 2,2'-bipyridine, dppz = dipyrido[3,2-a:2',3'-c]phenazine) (Ru-BD), was used as a spectroscopic probe for studying nucleic acid dynamics. The Ru-BD complex displays a long lifetime of over 100 ns and a molecular light switch property upon DNA binding due to shielding of its dppz ligand from water. To further show the usefulness of this luminophore (Ru-BD) for probing DNA dynamics, we examined its intensity and anisotropy decays when intercalated into supercoiled and linear pTZ18U plasmids using frequency-domain fluorometry with a light-emitting diode (LED) as the modulated light source. Compared to the supercoiled plasmids with an average intensity decay time of 120.8 ns at 25 degrees C, we obtained somewhat longer lifetimes for the linear plasmids ((tau) = 141.4 ns at 25 degrees C), suggesting a more efficient shielding from water by the linear plasmids. The anisotropy decay data also showed longer rotational correlation times for the linear plasmids (495 and 35 ns at 25 degrees C) as compared to the supercoiled plasmids (412 and 27 ns at 25 degrees C). The slow and fast rotational correlation times appear to be consistent with the bending and torsional motions of the plasmids, respectively. The anisotropy values were quite similar, although the values of the supercoiled plasmids were slightly higher in both the steady-state and anisotropy decay measurements. These results indicate that Ru-BD can be applied in the study of both bending and torsional dynamics of nucleic acids.