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Alternative Conformations (alternative + conformation)
Selected AbstractsDenaturation of replication protein A reveals an alternative conformation with intact domain structure and oligonucleotide binding activityPROTEIN SCIENCE, Issue 5 2004Jonathan E. Nuss Abstract Replication protein A (RPA) is a heterotrimeric, multidomain, single-stranded DNA-binding protein. Using spectroscopic methods and methylene carbene-based chemical modification methods, we have identified conformational intermediates in the denaturation pathway of RPA. Intrinsic protein fluorescence studies reveal unfolding profiles composed of multiple transitions, with midpoints at 1.5, 2.7, 4.2, and 5.3 M urea. CD profiles of RPA unfolding are characterized by a single transition. RPA is stabilized with respect to the CD-monitored transition when bound to a dA15 oligonucleotide. However, oligonucleotide binding appears to exert little, if any, effect on the first fluorescence transition. Methylene carbene chemical modification, coupled with MALDI-TOF mass spectrometry analysis, was also used to monitor unfolding of several specific RPA folds of the protein. The unfolding profiles of the individual structures are characterized by single transitions similar to the CD-monitored transition. Each fold, however, unravels with different individual characteristics, suggesting significant autonomy. Based on results from chemical modification and spectroscopic analyses, we conclude the initial transition observed in fluorescence experiments represents a change in the juxtaposition of binding folds with little unraveling of the domain structures. The second transition represents the unfolding of the majority of fold structure, and the third transition observed by fluorescence correlates with the dissociation of the 70- and 32-kD subunits. [source] Structure of the restriction,modification controller protein C.Esp1396IACTA CRYSTALLOGRAPHICA SECTION D, Issue 9 2009N. Ball The controller protein of the Esp1396I restriction,modification (R,M) system binds differentially to three distinct operator sequences upstream of the methyltransferase (M) and endonuclease (R) genes to regulate the timing of gene expression. The crystal structure of a complex of the protein with two adjacent operator DNA sequences has been reported; however, the structure of the free protein has not yet been determined. Here, the crystal structure of the free protein is reported, with seven dimers in the asymmetric unit. Two of the 14 monomers show an alternative conformation to the major conformer in which the side chains of residues 43,46 in the loop region flanking the DNA-recognition helix are displaced by up to 10,Å. It is proposed that the adoption of these two conformational states may play a role in DNA-sequence promiscuity. The two alternative conformations are also found in the R35A mutant structure, which is otherwise identical to the native protein. Comparison of the free and bound protein structures shows a 1.4,Å displacement of the recognition helices when the dimer is bound to its DNA target. [source] Consistent treatment of inter- and intramolecular polarization in molecular mechanics calculationsJOURNAL OF COMPUTATIONAL CHEMISTRY, Issue 16 2002Pengyu Ren Abstract A protocol is described for the treatment of molecular polarization in force field calculations. The resulting model is consistent in that both inter- and intramolecular polarization are handled within a single scheme. An analytical formula for removing intramolecular polarization from a set of atomic multipoles for an arbitrary static structure or conformation is given. With the help of the intramolecular polarization, these permanent atomic multipoles can then be applied in modeling alternative conformations of a molecule. Equipped with this simple technique, one can derive transferable electrostatic parameters for peptides and proteins using flexible model compounds such as dipeptides. The proposed procedure is tested for its ability to describe the electrostatic potential around various configurations of the N -methylacetamide dimer. The effect of different intramolecular polarization schemes on the accuracy of a force field model of the electrostatic potential of alanine dipeptide is investigated. A group-based scheme for including direct intramolecular polarization is shown to be most successful in accounting for the conformational dependence of electrostatic potentials. © 2002 Wiley Periodicals, Inc. J Comput Chem 23: 1497,1506, 2002 [source] Physical-chemical determinants of turn conformations in globular proteinsPROTEIN SCIENCE, Issue 8 2007Timothy O. Street Abstract Globular proteins are assemblies of ,-helices and ,-strands, interconnected by reverse turns and longer loops. Most short turns can be classified readily into a limited repertoire of discrete backbone conformations, but the physical,chemical determinants of these distinct conformational basins remain an open question. We investigated this question by exhaustive analysis of all backbone conformations accessible to short chain segments bracketed by either an ,-helix or a ,-strand (i.e., ,-segment-,, ,-segment-,, ,-segment-,, and ,-segment-,) in a nine-state model. We find that each of these four secondary structure environments imposes its own unique steric and hydrogen-bonding constraints on the intervening segment, resulting in a limited repertoire of conformations. In greater detail, an exhaustive set of conformations was generated for short backbone segments having reverse-turn chain topology and bracketed between elements of secondary structure. This set was filtered, and only clash-free, hydrogen-bond,satisfied conformers having reverse-turn topology were retained. The filtered set includes authentic turn conformations, observed in proteins of known structure, but little else. In particular, over 99% of the alternative conformations failed to satisfy at least one criterion and were excluded from the filtered set. Furthermore, almost all of the remaining alternative conformations have close tolerances that would be too tight to accommodate side chains longer than a single ,-carbon. These results provide a molecular explanation for the observation that reverse turns between elements of regular secondary can be classified into a small number of discrete conformations. [source] Structure analysis of endosialidase NF at 0.98,Å resolutionACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2010Eike C. Schulz Endosialidase NF (endoNF) is a bacteriophage-derived endosialidase that specifically degrades ,-2,8-linked polysialic acid. The structure of a new crystal form of endoNF in complex with sialic acid has been refined at 0.98,Å resolution. The 210,kDa homotrimeric multi-domain enzyme displays outstanding stability and resistance to SDS. Even at atomic resolution, only a minor fraction of side chains possess alternative conformations. However, multiple conformations of an active-site residue imply that it has an important catalytic function in the cleavage mechanism of polysialic acid. [source] Structure of the restriction,modification controller protein C.Esp1396IACTA CRYSTALLOGRAPHICA SECTION D, Issue 9 2009N. Ball The controller protein of the Esp1396I restriction,modification (R,M) system binds differentially to three distinct operator sequences upstream of the methyltransferase (M) and endonuclease (R) genes to regulate the timing of gene expression. The crystal structure of a complex of the protein with two adjacent operator DNA sequences has been reported; however, the structure of the free protein has not yet been determined. Here, the crystal structure of the free protein is reported, with seven dimers in the asymmetric unit. Two of the 14 monomers show an alternative conformation to the major conformer in which the side chains of residues 43,46 in the loop region flanking the DNA-recognition helix are displaced by up to 10,Å. It is proposed that the adoption of these two conformational states may play a role in DNA-sequence promiscuity. The two alternative conformations are also found in the R35A mutant structure, which is otherwise identical to the native protein. Comparison of the free and bound protein structures shows a 1.4,Å displacement of the recognition helices when the dimer is bound to its DNA target. [source] Modulation of activity by Arg407: structure of a fungal ,-1,2-mannosidase in complex with a substrate analogueACTA CRYSTALLOGRAPHICA SECTION D, Issue 3 2008Yuri D. Lobsanov Class I ,-mannosidases (glycoside hydrolase family GH47) play key roles in the maturation of N-glycans and the ER-associated degradation of unfolded glycoproteins. The 1.95,Å resolution structure of a fungal ,-1,2-mannosidase in complex with the substrate analogue methyl-,- d -lyxopyranosyl-(1,,2)-,- d -mannopyranoside (LM) shows the intact disaccharide spanning the ,1/+1 subsites, with the d -lyxoside ring in the ,1 subsite in the 1C4 chair conformation, and provides insight into the mechanism of catalysis. The absence of the C5, hydroxymethyl group on the d -lyxoside moiety results in the side chain of Arg407 adopting two alternative conformations: the minor one interacting with Asp375 and the major one interacting with both the d -lyxoside and the catalytic base Glu409, thus disrupting its function. Chemical modification of Asp375 has previously been shown to inactivate the enzyme. Taken together, the data suggest that Arg407, which belongs to the conserved sequence motif RPExxE, may act to modulate the activity of the enzyme. The proposed mechanism for modulating the activity is potentially a general mechanism for this superfamily. [source] The structure of a d(gcGAACgc) duplex containing two consecutive bulged A residues in both strands suggests a molecular switchACTA CRYSTALLOGRAPHICA SECTION D, Issue 6 2007Jiro Kondo In previous studies, it was reported that DNA fragments with the sequence d(gcGXYAgc) (where X = A or G and Y = A, T or G) form a stable base-intercalated duplex (Bi-duplex) in which the central X and Y residues are not involved in any base-pair interactions but are alternately stacked on each other between the two strands. To investigate the structural stability of the Bi-duplex, the crystal structure of d(gcGAACgc) with a point mutation at the sixth residue of the sequence, d(gcGAAAgc), has been determined. The two strands are associated in an antiparallel fashion to form two types of bulge-containing duplexes (Bc-duplexes), I and II, both of which are quite different from the Bi-duplex of the parent sequence. In both Bc-duplexes, three Watson,Crick G·C base pairs constitute the stem regions at the two ends. The A4 residues are bulged in to form a pair with the corresponding A4 residue of the opposite strand in either duplex. The A4·A4* pair formation is correlated to the orientations of the adjacent A5 residues. A remarkable difference between the two Bc-duplexes is seen at the A5 residue. In Bc-duplex I, it is flipped out and comes back to interact with the G3 residue. In Bc-duplex II, the A5 residue extends outwards to interact with the G7 residue of the neighbouring Bc-duplex I. These results indicate that trans sugar-edge/Hoogsteen (sheared-type) G3·A6* base pairs are essential in the formation of a Bi-duplex of d(gcGXYAgc). On the other hand, the alternative conformations of the internal loops containing two consecutive bulged A residues suggest molecular switching. [source] Crystal structure reveals two alternative conformations in the active site of ribonuclease Sa2ACTA CRYSTALLOGRAPHICA SECTION D, Issue 7 2004Jozef Three different strains of Streptomyces aureofaciens produce the homologous ribonucleases Sa, Sa2 and Sa3. The crystal structures of ribonuclease Sa (RNase Sa) and its complexes with mononucleotides have previously been reported at high resolution. Here, the structures of two crystal forms (I and II) of ribonuclease Sa2 (RNase Sa2) are presented at 1.8 and 1.5 Å resolution. The structures were determined by molecular replacement using the coordinates of RNase Sa as a search model and were refined to R factors of 17.5 and 15.0% and Rfree factors of 21.8 and 17.2%, respectively. The asymmetric unit of crystal form I contains three enzyme molecules, two of which have similar structures to those seen for ribonuclease Sa, with Tyr87 at the bottom of their active sites. In the third molecule, Tyr87 has moved substantially: the CA atom moves almost 5,Å and the OH of the side chain moves 10,Å, inserting itself into the active site of a neighbouring molecule at a similar position to that observed for the nucleotide base in RNase Sa complexes. The asymmetric unit of crystal form II contains two Sa2 molecules, both of which are similar to the usual Sa structures. In one molecule, two main-chain conformations were modelled in the ,-helix. Finally, a brief comparison is made between the conformations of the Sa2 molecules and those of 34 independent molecules taken from 20 structures of ribonuclease Sa and two independent molecules taken from two structures of ribonuclease Sa3 in various crystal forms. [source] The three-dimensional structure of cytosolic bovine retinal creatine kinaseACTA CRYSTALLOGRAPHICA SECTION D, Issue 2 2001Dominic Tisi Creatine kinase (CK) catalyses the reversible transfer of the phosphate moiety from phosphocreatine (PCr) to ADP, generating creatine and ATP. The crystal structure of a cytosolic brain-type creatine kinase is reported at 2.3,Å. The biological dimer sits on a crystallographic twofold axis. The N-terminal residues of both subunits come very close to the crystallographic twofold at the dimer interface. The electron density observed is consistent with two alternative conformations for the N-termini, as previously found for chicken brain-type creatine kinase. [source] | ||||||||||