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861.
Mazon H Marcillat O Forest E Vial C 《Protein science : a publication of the Protein Society》2004,13(2):476-486
Creatine kinase (CK) isoenzymes catalyse the reversible transfer of a phosphoryl group from ATP onto creatine. This reaction plays a very important role in the regulation of intracellular ATP concentrations in excitable tissues. CK isoenzymes are highly resistant to proteases in native conditions. To appreciate localized backbone dynamics, kinetics of amide hydrogen exchange with deuterium was measured by pulse-labeling the dimeric cytosolic muscle CK isoenzyme. Upon exchange, the protein was digested with pepsin, and the deuterium content of the resulting peptides was determined by liquid chromatography coupled to mass spectrometry (MS). The deuteration kinetics of 47 peptides identified by MS/MS and covering 96% of the CK backbone were analyzed. Four deuteration patterns have been recognized: The less deuterated peptides are located in the saddle-shaped core of CK, whereas most of the highly deuterated peptides are close to the surface and located around the entrance to the active site. Their exchange kinetics are discussed by comparison with the known secondary and tertiary structures of CK with the goal to reveal the conformational dynamics of the protein. Some of the observed dynamic motions may be linked to the conformational changes associated with substrate binding and catalytic mechanism. 相似文献
862.
Garcia-Viloca M Poulsen TD Truhlar DG Gao J 《Protein science : a publication of the Protein Society》2004,13(9):2341-2354
A subject of great practical importance that has not received much attention is the question of the sensitivity of molecular dynamics simulations to the initial X-ray structure used to set up the calculation. We have found two cases in which seemingly similar structures lead to quite different results, and in this article we present a detailed analysis of these cases. The first case is acyl-CoA dehydrogenase, and the chief difference of the two structures is attributed to a slight shift in a backbone carbonyl that causes a key residue (the proton-abstracting base) to be in a bad conformation for reaction. The second case is xylose isomerase, and the chief difference of the two structures appears to be the ligand sphere of a Mg2+ metal cofactor that plays an active role in catalysis. 相似文献
863.
Miranda FF Thórólfsson M Teigen K Sanchez-Ruiz JM Martínez A 《Protein science : a publication of the Protein Society》2004,13(5):1219-1226
Phosphorylation of phenylalanine hydroxylase (PAH) at Ser16 by cAMP-dependent protein kinase increases the basal activity of the enzyme and its resistance to tryptic proteolysis. The modeled structures of the full-length phosphorylated and unphosphorylated enzyme were subjected to molecular dynamics simulations, and we analyzed the energy of charge-charge interactions for individual ionizable residues in the final structures. These calculations showed that the conformational changes induced by incorporation of phosphate were localized and limited mostly to the region around the phosphoserine (Arg13-Asp17) and a region around the active site in the catalytic domain that includes residues involved in the binding of the iron and the substrate L-Phe (Arg270 and His285). The absence of a generalized conformational change was confirmed by differential scanning calorimetry, thermal-dependent circular dichroism, fluorescence spectroscopy, and limited chymotryptic proteolysis of the phosphorylated and unphosphorylated PAH. Our results explain the effect of phosphorylation of PAH on both the resistance to proteolysis specifically by trypsin-like enzymes and on the increase in catalytic efficiency. 相似文献
864.
Solvent effects on the conformational transition of a model polyalanine peptide 总被引:4,自引:0,他引:4 下载免费PDF全文
Nguyen HD Marchut AJ Hall CK 《Protein science : a publication of the Protein Society》2004,13(11):2909-2924
We have investigated the folding of polyalanine by combining discontinuous molecular dynamics simulation with our newly developed off-lattice intermediate-resolution protein model. The thermodynamics of a system containing a single Ac-KA(14)K-NH(2) molecule has been explored by using the replica exchange simulation method to map out the conformational transitions as a function of temperature. We have also explored the influence of solvent type on the folding process by varying the relative strength of the side-chain's hydrophobic interactions and backbone hydrogen bonding interactions. The peptide in our simulations tends to mimic real polyalanine in that it can exist in three distinct structural states: alpha-helix, beta-structures (including beta-hairpin and beta-sheet-like structures), and random coil, depending upon the solvent conditions. At low values of the hydrophobic interaction strength between nonpolar side-chains, the polyalanine peptide undergoes a relatively sharp transition between an alpha-helical conformation at low temperatures and a random-coil conformation at high temperatures. As the hydrophobic interaction strength increases, this transition shifts to higher temperatures. Increasing the hydrophobic interaction strength even further induces a second transition to a beta-hairpin, resulting in an alpha-helical conformation at low temperatures, a beta-hairpin at intermediate temperatures, and a random coil at high temperatures. At very high values of the hydrophobic interaction strength, polyalanines become beta-hairpins and beta-sheet-like structures at low temperatures and random coils at high temperatures. This study of the folding of a single polyalanine-based peptide sets the stage for a study of polyalanine aggregation in a forthcoming paper. 相似文献
865.
To investigate whether changes in land use and associated forest patch turnover affected genetic diversity and structure of the forest herb Primula elatior, historical data on landscape changes were combined with a population genetic analysis using dominant amplified fragment length polymorphism markers. Based on nine topographic maps, landscape history was reconstructed and forest patches were assigned to two age classes: young (less than 35 years) and old (more than 35 years). The level of differentiation among Primula populations in recently established patches was compared with the level of differentiation among populations in older patches. Genetic diversity was independent of population size (P > 0.05). Most genetic variation was present within populations. Within-population diversity levels tended to be higher for populations located in older forests compared with those for populations located in young forests (Hj = 0.297 and 0.285, respectively). Total gene diversity was also higher for old than for young populations (Ht = 0.2987 and 0.2828, respectively). The global fixation index FST averaged over loci was low, but significant. Populations in older patches were significantly more differentiated from each other than were populations in recently established patches and they showed significant isolation by distance. In contrast, no significant correlations between pairwise geographical distance and FST were found for populations in recently established patches. The location of young and old populations in the studied system and altered gene flow because of increased population density and decreased inter-patch distances between extant populations may explain the observed lower genetic differentiation in the younger populations. This study exemplifies the importance of incorporating data on historical landscape changes in population genetic research at the landscape scale. 相似文献
866.
Past and current gene flow in the selfing, wind-dispersed species Mycelis muralis in western Europe 总被引:2,自引:0,他引:2
Chauvet S van der Velde M Imbert E Guillemin ML Mayol M Riba M Smulders MJ Vosman B Ericson L Bijlsma R Giles BE 《Molecular ecology》2004,13(6):1391-1407
The distribution of genetic diversity in Mycelis muralis, or wall lettuce, was investigated at a European scale using 12 microsatellite markers to infer historical and contemporary forces from genetic patterns. Mycelis muralis has the potential for long-distance seed dispersal by wind, is mainly self-pollinated, and has patchily distributed populations, some of which may show metapopulation dynamics. A total of 359 individuals were sampled from 17 populations located in three regions, designated southern Europe (Spain and France), the Netherlands, and Sweden. At this within-region scale, contemporary evolutionary forces (selfing and metapopulation dynamics) are responsible for high differentiation between populations (0.34 < F(ST) < 0.60) but, contrary to expectation, levels of within-population diversity, estimated by Nei's unbiased expected heterozygosity (H(E)) (0.24 < H(E) < 0.68) or analyses of molecular variance (50% of the variation found within-populations), were not low. We suggest that the latter results, which are unusual in selfing species, arise from efficient seed dispersal that counteracts population turnover and thus maintains genetic diversity within populations. At the European scale, northern regions showed lower allelic richness (A = 2.38) than populations from southern Europe (A = 3.34). In light of postglacial colonization hypotheses, these results suggest that rare alleles may have been lost during recolonization northwards. Our results further suggest that mutation has contributed to genetic differentiation between southern and northern Europe, and that Sweden may have been colonized by dispersers originating from at least two different refugia. 相似文献
867.
The processes by which protein side chains reach equilibrium during a folding reaction are investigated using both lattice and all-atom simulations. We find that rates of side-chain relaxation exhibit a distribution over the protein structure, with the fastest relaxing side chains located in positions kinetically important for folding. Traversal of the major folding transition state corresponds to the freezing of a small number of side chains, belonging to the folding nucleus, whereas the rest of the protein proceeds toward equilibrium via backbone fluctuations around the native fold. The postnucleation processes by which side chains relax are characterized by very slow dynamics and many barrier crossings, and thus resemble the behavior of a glass. 相似文献
868.
Free-energy landscape of a chameleon sequence in explicit water and its inherent alpha/beta bifacial property 下载免费PDF全文
A sequence in yeast MATalpha2/MCM1/DNA complex that folds into alpha-helix or beta-hairpin depending on the surroundings has been known as "chameleon" sequence. We obtained the free-energy landscape of this sequence by using a generalized-ensemble method, multicanonical molecular dynamics simulation, to sample the conformational space. The system was expressed with an all-atom model in explicit water, and the initial conformation for the simulation was a random one. The free-energy landscape demonstrated that this sequence inherently has an ability to form either alpha or beta structure: The conformational distribution in the landscape consisted of two alpha-helical clusters with different packing patterns of hydrophobic residues, and four beta-hairpin clusters with different strand-strand interaction patterns. Narrow pathways connecting the clusters were found, and analysis on the pathways showed that a compact structure formed at the N-terminal root of the chameleon sequence controls the cluster-cluster transitions. The free-energy landscape indicates that a small conditional change induces alpha-beta transitions. Additional unfolding simulations done with replacing amino acids showed that the chameleon sequence has an advantage to form an alpha-helix. Current study may be useful to understand the mechanism of diseases resulting from abnormal chain folding, such as amyloid disease. 相似文献
869.
Motions and structural variability within toxins: implication for their use as scaffolds for protein engineering 下载免费PDF全文
Gilquin B Bourgoin M Ménez R Le Du MH Servent D Zinn-Justin S Ménez A 《Protein science : a publication of the Protein Society》2003,12(2):266-277
Animal toxins are small proteins built on the basis of a few disulfide bonded frameworks. Because of their high variability in sequence and biologic function, these proteins are now used as templates for protein engineering. Here we report the extensive characterization of the structure and dynamics of two toxin folds, the "three-finger" fold and the short alpha/beta scorpion fold found in snake and scorpion venoms, respectively. These two folds have a very different architecture; the short alpha/beta scorpion fold is highly compact, whereas the "three-finger" fold is a beta structure presenting large flexible loops. First, the crystal structure of the snake toxin alpha was solved at 1.8-A resolution. Then, long molecular dynamics simulations (10 ns) in water boxes of the snake toxin alpha and the scorpion charybdotoxin were performed, starting either from the crystal or the solution structure. For both proteins, the crystal structure is stabilized by more hydrogen bonds than the solution structure, and the trajectory starting from the X-ray structure is more stable than the trajectory started from the NMR structure. The trajectories started from the X-ray structure are in agreement with the experimental NMR and X-ray data about the protein dynamics. Both proteins exhibit fast motions with an amplitude correlated to their secondary structure. In contrast, slower motions are essentially only observed in toxin alpha. The regions submitted to rare motions during the simulations are those that exhibit millisecond time-scale motions. Lastly, the structural variations within each fold family are described. The localization and the amplitude of these variations suggest that the regions presenting large-scale motions should be those tolerant to large insertions or deletions. 相似文献
870.
Backbone dynamics of the human MIA protein studied by (15)N NMR relaxation: implications for extended interactions of SH3 domains 总被引:1,自引:0,他引:1 下载免费PDF全文
Stoll R Renner C Buettner R Voelter W Bosserhoff AK Holak TA 《Protein science : a publication of the Protein Society》2003,12(3):510-519
The melanoma inhibitory activity (MIA) protein is a clinically valuable marker in patients with malignant melanoma as enhanced values diagnose metastatic melanoma stages III and IV. Here, we report the backbone dynamics of human MIA studied by (15)N NMR relaxation experiments. The folded core of human MIA is found to be rigid, but several loops connecting beta-sheets, such as the RT-loop for example, display increased mobility on picosecond to nanosecond time scales. One of the most important dynamic features is the pronounced flexibility of the distal loop, comprising residues Asp 68 to Ala 75, where motions on time scales up to milliseconds occur. Further, significant exchange contributions are observed for residues of the canonical binding site of SH3 domains including the RT-loop, the n-Src loop, for the loop comprising residues 13 to 19, which we refer to as the"disulfide loop", in part for the distal loop, and the carboxyl terminus of human MIA. The functional importance of this dynamic behavior is discussed with respect to the biological activity of several point mutations of human MIA. The results of this study suggest that the MIA protein and the recently identified highly homologous fibrocyte-derived protein (FDP)/MIA-like (MIAL) constitute a new family of secreted proteins that adopt an SH3 domain-like fold in solution with expanded ligand interactions. 相似文献