Showing papers on "Cooperativity published in 1989"
1,357 citations
TL;DR: A cooperative quinolone-DNA binding model for the inhibition of DNA gyrase and the unique self-association phenomenon (from which the cooperativity is derived) of the drug molecules to fit the binding pocket with a high degree of flexibility is proposed.
Abstract: We have proposed a cooperative quinolone-DNA binding model for the inhibition of DNA gyrase. The essential feature of the model is that bound gyrase induces a specific quinolone binding site in the relaxed DNA substrate in the presence of ATP. The binding affinity and specificity are derived from two unique and equally important functional features: the specific conformation of the proposed single-stranded DNA pocket induced by the enzyme and the unique self-association phenomenon (from which the cooperativity is derived) of the drug molecules to fit the binding pocket with a high degree of flexibility. Supporting evidence for and implications of this model are provided.
412 citations
TL;DR: By exposing the individual contributions, the free energy simulation provides new insights into the origin of thermodynamic changes in mutant proteins and demonstrates the role of effects beyond those usually considered in structural analyses.
Abstract: A molecular dynamics simulation method is used to determine the contributions of individual amino acid residues and solvent molecules to free energy changes in proteins. Its application to the hemoglobin interface mutant Asp G1(99) beta----Ala shows that some of the contributions to the difference in the free energy of cooperativity are as large as 60 kilocalories (kcal) per mole. Since the overall free energy change is only -5.5 kcal/mole (versus the experimental value of -3.4 kcal/mole), essential elements of the thermodynamics are hidden in the measured results. By exposing the individual contributions, the free energy simulation provides new insights into the origin of thermodynamic changes in mutant proteins and demonstrates the role of effects beyond those usually considered in structural analyses.
248 citations
TL;DR: Research from various laboratories gives support to the binding change mechanism in which energy from proton translocation serves principally to promote release of tightly bound ATP, with sequential participation of three catalytic sites.
Abstract: An overview of research in the field of bioenergetics that led to the development of the binding change mechanism for ATP synthesis is presented, with emphasis on research from the author's laboratory. The text follows closely the Rose Award Lecture given at the 1989 meeting of the American Society for Biochemistry and Molecular Biology. Remarkable advances have revealed that the ubiquitous membrane-bound ATP synthase has unusual composition and properties. The enzyme complex has 1, 2, 3, or 9-12 copies of eight or more protein subunits. The catalytic sites are located on three copies of an approximately 55-kDa subunit. It has the strongest positive catalytic cooperativity known for any enzyme. Examples are given of selected experimental results that have provided insights into its mechanism. These include demonstration of the characteristics, location, and function of catalytic and noncatalytic adenine nucleotide binding sites and the incisive information provided by measurement of phosphate oxygen exchanges and distribution of 18(O) in ATP or Pi formed by catalysis. Research from various laboratories gives support to the binding change mechanism in which energy from proton translocation serves principally to promote release of tightly bound ATP, with sequential participation of three catalytic sites. Some speculative suggestions about a rotational catalysis and about the different forms assumed by the ATPase are included.
242 citations
TL;DR: The structure and properties of VirE2-ssDNA complexes predict possible functions in Agrobacterium virulence to protect the T-strands from cellular nucleases and facilitate transfer of the T -strands through bacterial membranes possibly by specific interaction with putative membrane pores formed in plant-induced Agrobacteria cells.
Abstract: Induction of Agrobacterium tumefaciens vir gene expression by wounded plant cells results in production of a free transferable single-stranded (ss) copy of T-DNA, the T-strand. One of the Vir proteins, the VirE2 polypeptide, is a ssDNA-binding protein. In the present work, interaction of nopaline-specific VirE2 protein (Mr 69,000) with ssDNA was studied by using nitrocellulose filter binding, gel retardation, and electron microscopy techniques. The VirE2 protein was found to bind to ssDNA molecules with strong cooperativity, forming VirE2-ssDNA complexes with a binding site of 28-30 nucleotides. The VirE2-ssDNA complexes are stable at high salt concentrations and resistant to exonucleolytic activity. When examined under the electron microscope, the VirE2 protein converted collapsed free ssDNA molecules into unfolded and extended structures. The structure and properties of VirE2-ssDNA complexes predict possible functions in Agrobacterium virulence to (i) protect the T-strands from cellular nucleases and (ii) facilitate transfer of the T-strands through bacterial membranes possibly by specific interaction with putative membrane pores formed in plant-induced Agrobacterium cells.
211 citations
TL;DR: It is demonstrated that pre-tRNA(Leu-3) responds to Mg2+ depletion in the same manner as detected by the appearance of highly sensitive RNase cleavage sites in the D and T psi C loops.
Abstract: To make strong statements about possible tertiary structure or the relative stability of regions of secondary structure, the structure-probing experiments must go further than single-hit reactions. Some elements of the environment of the RNA molecule must be altered systematically. Knowledge of the effects of ions or other interacting factors on the activity or physical parameters (e.g., NMR and melting cooperativity) of the RNA help in experimental design. For example, the copious work on tRNA(Phe) compared the crystal and solution structures and allowed the direct correlation of Mg2+ stabilization of the tertiary structure of that molecule. Figure 3 demonstrates that pre-tRNA(Leu-3) responds to Mg2+ depletion in the same manner as detected by the appearance of highly sensitive RNase cleavage sites in the D and T psi C loops. Similar experiments titrating polyamine concentrations suggested that secondary structure was more efficiently stabilized by polyamines than by Mg2+. The variation of Mg2+ concentrations has been used to gain additional information about other RNA structures. Others have used protein-RNA interactions to approach the question of the functional structure of a RNA (for examples, see Ref. 3). Thus, the ideal parameters to choose would be those known to affect the function of the RNA. The variation of Mg2+ and polyamine concentrations would minimally suggest regions of greater or lesser secondary or tertiary structure stability.
191 citations
TL;DR: The results suggest that the drug binds to unpaired bases via hydrogen bonding and not via ring stacking with DNA bases, and binding saturation curves of this type are highly cooperative (with Hill constant greater than 4).
Abstract: Although the functional target of quinolone antibacterials such as nalidixic acid and norfloxacin has been identified as the enzyme DNA gyrase, the direct binding site of the drug is the DNA molecule [Shen, L. L., & Pernet, A. G. (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 307-311]. As described in this paper, binding specificity and cooperativity of quinolones to DNA were further investigated with the use of a variety of DNA species of different structures and different base compositions. Results show that the drug binding specificity is controlled and determined largely by the DNA structure. The drug binds weakly and demonstrates no base preference when DNA strands are paired. The drug binds with much greater affinity when the strands are separated, and consequently, binding preference emerges: it binds better to poly(G) and poly(dG) over their counterparts including poly(dI). The results suggest that the drug binds to unpaired bases via hydrogen bonding and not via ring stacking with DNA bases. The weak binding to relaxed double-stranded DNA and the stronger binding to single-stranded DNA are both nonspecific as they do not demonstrate binding saturation and cooperativity. The specific type of binding, initially demonstrated in our previous publication with the supercoiled DNA and more recently with complex formed between linear DNA and DNA gyrase [Shen, L. L., Kohlbrenner, W. E., Weigl, D., & Baranowski, J. (1989) J. Biol. Chem. (in press)], occurs near the drug's supercoiling inhibition concentration. As shown in this paper, binding saturation curves of this type are highly cooperative (with Hill constant greater than 4).(ABSTRACT TRUNCATED AT 250 WORDS)
180 citations
TL;DR: Phosphofructokinase from Bacillus stearothermophilus shows cooperative kinetics with respect to the substrate fructose-6-phosphate, allosteric activation by ADP, and inhibition by phosphoenolpyruvate, which suggests that a rearrangement of the subunits could explain the cooperativity of substrate binding.
Abstract: Phosphofructokinase from Bacillus stearothermophilus shows cooperative kinetics with respect to the substrate fructose-6-phosphate (F6P), allosteric activation by ADP, and inhibition by phosphoenolpyruvate. The crystal structure of the active conformation of the enzyme has been solved to 2.4 A resolution, and three ligand-binding sites have been located. Two of these form the active site and bind the substrates F6P and ATP. The third site binds both allosteric activator and inhibitor. The complex of the enzyme with F6P and ADP has been partly refined at 2.4 A resolution, and a model of ATP has been built into the active site by using the refined model of ADP and a 6 A resolution map of bound 5'-adenylylimidodiphosphate (AMPPNP). The $\gamma$ -phosphate of ATP is close to the 1-hydroxyl of F6P, in a suitable position for in-line phosphoryl transfer. The binding of the phosphate of F6P involves two arginines from a neighbouring subunit in the tetramer, which suggests that a rearrangement of the subunits could explain the cooperativity of substrate binding. The activator ADP is also bound by residues from two subunits.
169 citations
TL;DR: These studies indicate that protein B23 binds cooperatively with high affinity for single-stranded nucleic acids and exhibits RNA helix destabilizing activity, which may be related to its role in ribosome assembly.
Abstract: The interaction of eukaryotic nucleolar phosphoprotein B23 with nucleic acids was examined by gel retardation and filter binding assays, by fluorescence techniques, and by circular dichroism All studies utilized protein prepared under native conditions by a newly developed purification procedure Electrophoretic gel mobility shift assays with phage M13 DNA suggested that protein B23 is a single-stranded nucleic acid binding protein This was confirmed in competition binding assays with native or heat-denatured linearized plasmid pUC18 DNA where the protein showed a marked preference for the denatured form In other competition assays, there was no apparent preference for single-stranded synthetic ribo- versus deoxyribonucleotides Equilibrium binding with poly(riboethenoadenylic acid) indicated cooperative ligand binding with a protein binding site size of 11 nucleotides and an apparent binding constant (K omega) of 5 x 10(7) M-1 which includes an intrinsic binding constant (K) of 63 x 10(4) M-1 and a cooperativity factor (omega) of 800 In circular dichroism (CD) studies, protein B23, when combined with the single-stranded synthetic nucleic acids poly(rA) and poly(rC), effected a decrease in ellipticity and a shift of the positive peak at 260-270 nm toward higher wavelengths, indicating helix destabilizing activity No CD changes were seen with double-stranded poly(dAdT) The change in ellipticity of poly(rA) was sigmoidal upon addition of protein, confirming the cooperative behavior seen with fluorescence methods These studies indicate that protein B23 binds cooperatively with high affinity for single-stranded nucleic acids and exhibits RNA helix destabilizing activity These features may be related to its role in ribosome assembly
166 citations
TL;DR: In this article, the time dependence of pair-interaction data obtained from a molecular dynamics simulation of 216 ST2 [F. H. Stillinger and A. Rahman, J. Chem. Phys. 60, 1545 (1974)] water molecules at 280 K was analyzed.
Abstract: Hydrogen bonding in water systems is investigated by introducing a new method to analyze the time dependence of pair‐interaction data obtained from a molecular dynamics simulation of 216 ST2 [F. H. Stillinger and A. Rahman, J. Chem. Phys. 60, 1545 (1974)] water molecules at 280 K. This approach avoids the use of cutoff values and yields a more realistic bond population, whose distributions of geometric and energetic properties are reported as a function of the bond lifetimes. For the fraction of long‐lived bonds, correlation among bond stability, molecular mobility, and local structure is elecited. Percolation analysis of HB network evidences cooperativity in the spatial distribution of bonds, which does not originate from proton polarizability of HB and/or from many‐body terms of the interaction potentials since a rigid water model and pair potentials are used. These features can play a role in the anomalous properties of liquid water.
157 citations
TL;DR: Kinetic studies of the phosphoenzyme intermediates of site-specific mutants were used to examine the role of Gly233 in the reaction mechanism of the sarcoplasmic reticulum Ca2(+)-ATPase, suggesting the lack of a role of this region of the peptide in formation of Ca2 (+)-binding sites.
TL;DR: Application of the principle of detailed balance shows that with only one ligand binding site the receptor will not be able to derive enough energy from acetylcholine binding to drive a sufficiently large change in the channel conformational equilibrium, which provides a rationale for the existence of a second binding site.
Abstract: The energetics and kinetics of activation of the acetylcholine receptor are evaluated in the context of optimizing rapid synaptic transmission. Physiological needs are used as the basis for estimating optimal values for the closed-to-open channel equilibrium constants of the liganded and unliganded receptor. An estimate is made of the maximum energy that can be derived from the binding of acetylcholine to a perfectly designed receptor binding site. Application of the principle of detailed balance shows that with only one ligand binding site the receptor will not be able to derive enough energy from acetylcholine binding to drive a sufficiently large change in the channel conformational equilibrium. This then provides a rationale for the existence of a second binding site, rather than the often invoked advantage of cooperativity. With two binding sites there is a considerable excess of binding energy and consequently considerable flexibility in how binding energy can be utilized. It is shown that the receptor must have at least one binding site that binds acetylcholine weakly when the channel is closed. This is essential to rapid response termination. However, making the other binding site bind more tightly can enhance and accelerate the activation of the receptor. To optimize both response activation and termination the best solution is to make the two binding sites different in their binding affinities. This qualitatively reproduces an experimental observation.
TL;DR: It is proposed that Ca2+ and acidic phospholipids cause the protein to undergo a conformation change revealing multiple phosphatidylserine binding sites and resulting in the highly cooperative and specific interaction of protein kinase C with phosphatilserine.
TL;DR: Cooperativity in OTF-2 binding is closely mirrored by functional cooperation between the heptamer and octamer elements in activating transcription from the heavy-chain promoter in vitro.
Abstract: Immunoglobulin heavy-chain gene promoters contain two conserved upstream sequence elements, octamer and heptamer, both of which are required for normal cell type-specific promoter function in vivo. The octamer sequence motif 5'-ATGCAAAT-3', and its precise inverse, are strongly conserved in heavy- and light-chain gene promoters and are important determinants for the lymphoid-specific function of these promoters and of the heavy-chain enhancer. The heptameric sequence element with the consensus 5'-CTCATGA-3' (refs 3 and 4) is also required in addition to the octamer for full lymphoid-specific activity of heavy-chain promoters. Although these two elements have no sequence similarity, they are both recognized in vitro by the ubiquitous octamer transcription factor OTF-1 (reviewed refs 13 and 14) and the lymphoid-specific OTF-2 (reviewed in refs 15 and 16). Here we show that purified OTF-2 binds cooperatively to the immunoglobulin heptamer and octamer elements so that interaction with the octamer element facilitates binding of OTF-2 to the heptamer motif. More important, cooperativity in OTF-2 binding is closely mirrored by functional cooperation between the heptamer and octamer elements in activating transcription from the heavy-chain promoter in vitro.
TL;DR: The alpha 3 beta 3 complex was reconstituted from alpha and beta subunits of the thermophilic bacterium PS3 F1-ATPase and showed that the essential structure for eliciting the ATPase activity of F 1- ATPase is trimeric alpha beta pairs and that the kinetic cooperativity of the F1/2 subunit is an inherent property of this trimeric structure but is not due to the presence of single-copy subunits.
Abstract: The alpha 3 beta 3 complex was reconstituted from alpha and beta subunits of the thermophilic bacterium PS3 F1-ATPase (TF1) and then isolated. It is less stable at high and low temperatures than TF1, and the complex dissociates into subunits during native polyacrylamide gel electrophoresis. The alpha 3 beta 3 complex has about 20% of the ATPase activity of TF1. Its enzymic properties are similar to those of the native TF1, exhibiting similar cooperative kinetics as a function of ATP concentration, similar substrate specificity for nucleotide triphosphates, and the presence of two peaks in its temperature-activity profile. Differing from TF1, the ATPase activity of the alpha 3 beta 3 complex is insensitive to N3- inhibition, its divalent cation specificity is less stringent, and its optimum pH shifts to the alkaline side. The addition of the gamma subunit to the alpha 3 beta 3 complex leads to the formation of the alpha 3 beta 3 gamma complex, indicating that the alpha 3 beta 3 complex is an intermediate in the process of assembly of the holoenzyme from each subunit. These results definitely show that the essential structure for eliciting the ATPase activity of F1-ATPase is trimeric alpha beta pairs and that the kinetic cooperativity of the F1-ATPase is an inherent property of this trimeric structure but is not due to the presence of single-copy subunits. In this sense, the alpha 3 beta 3 complex is the catalytic core of F1-ATPase.
TL;DR: All four Adair constants could be determined for alpha alpha Hb under most conditions, but a3 could not be resolved for HbAo without constraining a4, suggesting that the cross-link stabilizes triply ligated intermediates of hemoglobin.
TL;DR: The cooperative interaction between Cu and ACE1 allows the cell to respond to a small change in metal concentration by a large change in gene expression, suggesting the effect of the metal on DNA binding is primarily kinetic rather than thermodynamic.
Abstract: Cu ions activate yeast metallothionein gene transcription by altering the conformation and DNA-binding activity of the ACE1 transcription factor. We show that this conformational switch occurs in an all-or-none highly cooperative fashion (Hill coefficient = 4). Analysis of the subunit composition of ACE1 bound to DNA indicates that cooperativity results from the binding of multiple Cu(I) ions to the cysteine-rich DNA-binding domain. Surprisingly, DNA has little effect on the interaction between Cu(I) and ACE1 as assayed by partial proteolysis; this suggests that the effect of the metal on DNA binding is primarily kinetic rather than thermodynamic. Although Ag(I) also activates ACE1, it acts less cooperatively than the smaller Cu(I) ion and the resulting metalloprotein has a reduced affinity for DNA. The cooperative interaction between Cu and ACE1 allows the cell to respond to a small change in metal concentration by a large change in gene expression.
TL;DR: This model indicates that oligomeric ligands (substrates) interact through cooperativity of their subunits at the extended binding site of cellobiohydrolase II, indicating 'transition-state'-type binding for these ligands at subsite C.
Abstract: Binding onto cellobiohydrolase II from Trichoderma reesei of glucose, cellobiose, cellotriose, derivatized and analogous compounds, is monitored by protein-difference-absorption spectroscopy and by titration of ligand fluorescence, either at equilibrium or by the stopped-flow technique. The data complete earlier results [van Tilbeurgh, H., Pettersson, L. G., Bhikhabhai, R., De Boeck, H. and Claeyssens, M. (1985) Eur. J. Biochem. 148, 329-334] indicating an extended active center, with putative subsites ABCD. Subsite A specifically complexes with beta-D-glucosides and D-glucose; at 25 degrees C the latter influences the concomitant binding of other ligands at neighbouring sites. For several ligands this cooperative effect for binding (at 0.33 M glucose and temperature range 4-37 degrees C) was characterized by a substantial increase of the enthalpic term (delta delta H = -35 kJ mol-1). Glucose (0.33 M) decreases the association and dissociation rate parameters of 4-methylumbelliferyl beta-D-cellobioside by one order of magnitude: k+ = (3.6 +/- 0.5) x 10(-5) M-1 s-1 versus (5.1 +/- 0.1) x 10(-6) M-1 s-1 (in the absence of glucose) and k- = (1.3 +/- 0.1) s-1 versus (14.0 +/- 0.3) s-1. As deduced from substrate-specificity studies and inhibition experiments, subsite B interacts with terminal non-reducing glucopyranosyl residues of oligomeric ligands and substrates, whereas catalytic (hydrolytic) cleavage occurs between C and D. Association constants 10-100 times higher than those for cellobiose or its glycosides were observed for D-glucopyranosyl-(1----4)-beta-D-xylopyranose and cellobionolactone derivatives, suggesting 'transition-state'-type binding for these ligands at subsite C. Although subsite D can accomodate a bulky chromophoric group (MeUmb) its preference for a glucosyl residue is reflected in the lower binding enthalpy of cellotriose (-34 kJ mol-1) as compared to cellobiose (-28.3 kJ mol-1) and MeUmb(Glc)2 (-11.6 kJ mol-1). This model indicates that oligomeric ligands (substrates) interact through cooperativity of their subunits at the extended binding site of cellobiohydrolase II.
TL;DR: In this paper, a functional group probing capability of FTIR together with X-ray and molecular modelling techniques was used to investigate the dynamic properties, conformational structure and phase behaviour of deep rough bacterial lipopolysaccharide (Re-LPS) have been investigated.
TL;DR: Three independent methods establish that enzyme concentration-dependent oligomerization is a mechanism of activation of the erythrocyte Ca2+-ATPase, which is independent of the previously described activation by calmodulin.
TL;DR: Analysis of the data using the two-state model of Hill et al. suggests that removal of TM overlap abolishes nearest neighbor interactions, while the concerted change of the state of 7 actins in a functional unit can account for the residual cooperativity.
TL;DR: The general secular equation for three‐state lattice models for the cooperative binding of large ligands to a one‐dimensional lattice is presented and a closed‐form expression for the isotherm is obtained that can be used with all values of the cooperativity parameter ω(0 < ω < ∞) thus eliminating the need for multiple equations.
Abstract: We present the general secular equation for three-state lattice models for the cooperative binding of large ligands to a one-dimensional lattice. In addition, a closed-form expression for the isotherm is also obtained, that can be used with all values of the cooperativity parameter omega(0 less than omega less than infinity) thus eliminating the need for multiple equations.
TL;DR: The crystal structure of the cooperative dimeric hemoglobin from the arcid clam, Scapharca inaequivalvis, has been determined in the carbonmonoxy state and the details of an assemblage of two identical myoglobin-like subunits that is radically different from vertebrate hemoglobins are revealed.
TL;DR: The infection-specific E2F was shown to bind cooperatively to a pair of E2f sites found upstream of the Ad2 early region 2 mRNA cap site, which was sensitive to the spacing between the sites and their relative orientation.
Abstract: E2F is a sequence-specific DNA-binding factor which binds to sites that occur in pairs upstream of the adenovirus E1A and E2 early transcriptional start sites. Substantial quantities of E2F activity were found in uninfected-cell extracts, and there was a modest increase in E2F activity during an adenovirus type 5 (Ad5) infection. In uninfected cells, E2F was found to exist in multiple forms that could be separated chromatographically. Extracts prepared at 24 h after Ad5 infection contained a new form of E2F. This infection-specific form may have been a modified version of one of the forms present in uninfected cells. The infection-specific E2F was shown to bind cooperatively to a pair of E2F sites found upstream of the Ad2 early region 2 mRNA cap site. This binding was sensitive to the spacing between the sites and their relative orientation. In contrast, E2F binding in uninfected-cell extracts was unaffected by changes in orientation and spacing, consistent with very low cooperativity or independent binding.
TL;DR: The Arg 28/Lys 139 double replacement changed the equilibrium folding model from two state to multistate and showed evidence for interaction in one of the two kinetic phases detected in both unfolding and refolding reactions.
Abstract: To test the possibility that long-range interactions might influence the folding and stability of dihydrofolate reductase, a series of single and double mutations at positions 28 and 139 were constructed and their urea-induced unfolding reactions studied by absorbance and circular dichroism spectroscopy. The alpha carbons of the two side chains are separated by 15 A in the native conformation. The replacement of Leu 28 by Arg and of Glu 139 by Gln resulted in additive effects on both kinetic and equilibrium properties of the reversible unfolding transition; no evidence for interaction was obtained. In contrast, the Arg 28/Lys 139 double replacement changed the equilibrium folding model from two state to multistate and showed evidence for interaction in one of the two kinetic phases detected in both unfolding and refolding reactions. The results can be explained in terms of a long-range, repulsive electrostatic interaction between the cationic side chains at these two positions.
TL;DR: It is proposed that the novel Ag18-MT complex forms with a structure that involves a well defined three-dimensional structure, in the same manner as that recently reported for the Hg18- MT complex.
TL;DR: Replacement of the amino-terminal region of the E2 trans- activation domain with a synthetic amphipathic helix partially restored the trans-activation function; however, it did not result in a molecule that exhibited cooperativity between neighboring E2 DNA-binding sites.
Abstract: The papilloma virus E2 transcriptional trans-activator is representative of a class of transcriptional modulators that activate transcription through direct binding to cis-acting DNA sequences. In this study we measured the capacity for this mammalian virus factor to function in Saccharomyces cerevisiae. When expressed in the yeast, the bovine papilloma virus E2 trans-activator could stimulate transcription from a yeast promoter having E2 DNA-binding sites present in cis. Whereas a single E2 DNA-binding site was sufficient for trans-activation, a strong cooperative effect was observed with two E2 DNA-binding sites. The level of trans-activation was dependent on the position of the E2 DNA-binding sites in relation to the yeast promoter, with the maximal effect demonstrated when the binding sites were positioned upstream. Deleted E2 proteins, lacking part of the trans-activation or DNA-binding domains, failed to activate transcription in yeast, similar to their behavior in mammalian cells. Replacement of the amino-terminal region of the E2 trans-activation domain with a synthetic amphipathic helix partially restored the trans-activation function; however, it did not result in a molecule that exhibited cooperativity between neighboring E2 DNA-binding sites.
TL;DR: A method to analyze the DNA binding properties of RecA protein is developed to take into account the protein self-association reaction and is applied to reanalyze the interaction with chemically modified single-stranded DNA (epsilon-DNA).
TL;DR: The crystal structure of glycogen phosphorylase a complexed with its substrates, orthophosphate and maltopentaose, has been determined and refined and conformational changes in the core of the interface are correlated with those observed on covalent activation by phosphorylation at Ser14.
Abstract: The crystal structure of glycogen phosphorylase a complexed with its substrates, orthophosphate and maltopentaose, has been determined and refined at a resolution of 2.8 angstroms. With oligosaccaride bound at the glycogen storage site, the phosphate ion binds at the catalytic site and causes the regulatory and catalytic domains to separate with the loss of stabilizing interactions between them. Homotropic cooperativity between the active sites of the allosteric dimer results from rearrangements in isologous contacts between symmetry-related helices in the subunit interface. The conformational changes in the core of the interface are correlated with those observed on covalent activation by phosphorylation at Ser14 (phosphorylase b----a).