Proteomics and Models for Enzyme Cooperativity

Koshland, Daniel E.; Hamadani, Kambiz M.

doi:10.1074/jbc.r200014200

Cited by 144 publications

(132 citation statements)

References 90 publications

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“…Interestingly, Pyrococcus furiosis MBP binds maltotriose with a 20-fold greater affinity than E. coli MBP (32). The results from the present study suggest that this loop may have evolved specifically to function as a modulator of ligand binding affinity: mutations in the loop, which are relatively easy to accommodate, alter the affinity of the protein for its ligand by changing its conformational equilibria, and the loop therefore provides a mechanism for natural selection to alter ligand binding affinity without affecting specificity (33).…”

Section: High Resolution Analysis Of Structural Changes In Mbp-del Anmentioning

confidence: 70%

Insights into the Conformational Equilibria of Maltose-binding Protein by Analysis of High Affinity Mutants

Telmer

Shilton

2003

Journal of Biological Chemistry

110

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ATP binding cassette transporters couple ATP hydrolysis to the transmembrane transport of a diverse range of compounds. Members of the ATP binding cassette transporter superfamily are characterized by two membrane-integral domains that each contain 6 or more membrane spanning helices, but are otherwise poorly conserved, and two peripheral ATP binding cassette domains that display sequence conservation across the entire superfamily (1). In addition to the membrane complex, ATP binding cassette systems that catalyze nutrient uptake have primary receptors (binding proteins) that serve two functions: they provide a high affinity binding site for the transported molecule and they regulate the ATPase activity of the integral membrane complex.We are interested in the function of the primary receptors in the transport process. As a group, these proteins have been intensively studied by x-ray crystallography and other biophysical techniques (for a review, see Ref.2). They typically contain two domains separated by a hinge region; the substrate binds in the cleft between the two domains, and the protein undergoes a large conformational change, leading to closure of the cleft. With respect to the maltose transport system, domain closure in the binding protein is thought to be the first step toward molecular shape recognition by the membrane complex (3, 4), although it has been shown that both substrate-loaded and substrate-free binding proteins have a role in the transport cycle (5-8). The association and dissociation of the substrate, and attendant conformational changes in the binding protein (MBP), 1 may have direct effects on transport kinetics and regulation of ATP hydrolysis by MalFGK 2 . To investigate the role of binding protein affinity on the transport process, our goal was to engineer MBP molecules with greater affinity for maltose, without changing residues in either the maltose binding site or in regions thought to interact with MalFGK 2 .Crystal structures of MBP in both the closed and open conformations have been solved (9, 10), and they show that binding of maltose results in a large conformational change of the protein, bringing the two domains together such that the substrate is buried inside the cleft. In solution, unliganded MBP is in the open conformation (11); however, there is no obvious energetic barrier to closure of the ligand binding cleft, either in the hinge or in the interface surrounding the ligand binding site. Rather, an interface on the opposite side of the hinge from the ligand binding site appears to maintain the protein in an * This work was supported by Natural Sciences and Engineering Research Council Grant 21749-1999 (to B. H. S.); the macromolecular x-ray facility at the University of Western Ontario was financed with grants from the Canada Foundation for Innovation, Ontario Challenge Fund, and Western's Academic Development Fund; fluorescence and SPR studies were carried out at the University of Western Ontario Biomolecular Interactions and Conformations Facility, supported by a Mu...

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Section: High Resolution Analysis Of Structural Changes In Mbp-del Anmentioning

confidence: 70%

Insights into the Conformational Equilibria of Maltose-binding Protein by Analysis of High Affinity Mutants

Telmer

Shilton

2003

Journal of Biological Chemistry

110

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“…Negative cooperativity, on the other hand, spreads out this response over a wide concentration range. Interestingly, a recent survey of known cooperative enzymes finds that positive and negative cooperativity arise in the literature with approximately equal frequency (29). The consequence of negative cooperativity, in the PrP octarepeat domain, is that the protein transitions from 10 to 90% copper occupation over an extremely wide concentration range from 0.1 nM to 90 μM.…”

Section: Discussionmentioning

confidence: 99%

The Affinity of Copper Binding to the Prion Protein Octarepeat Domain: Evidence for Negative Cooperativity

2006

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The prion protein (PrP) binds Cu 2+ in its N-terminal octarepeat domain, composed of four or more tandem PHGGGWGQ segments. Previous work from our laboratory demonstrates that copper interacts with the octarepeat domain through three distinct coordination modes at pH 7.4, depending upon the precise ratio of Cu 2+ to protein. Here, we apply both electron paramagnetic resonance (EPR) and fluorescence quenching to determine the copper affinity for each of these modes. At low copper occupancy, which favors multiple His coordination, the octarepeat domain binds Cu 2+ with a dissociation constant of 0.10 (±0.08) nM. In contrast, high copper occupancy, involving coordination through deprotonated amide nitrogens, exhibits a weaker affinity characterized by dissociation constants in the range of 7.0-12.0 μM. Decomposition of the EPR spectra reveals the proportions of all coordination species throughout the copper concentration range and identifies significant populations of intermediates, consistent with negative cooperativity. At most copper concentrations, the Hill coefficient is less than 1.0 and approximately 0.7 at half copper occupancy. These findings demonstrate that the octarepeat domain is responsive to a remarkably wide copper concentration range covering approximately 5 orders of magnitude. Consideration of these findings, along with the demonstrated ability of the protein to quench copper redox activity at high occupancy, suggests that PrP may function to protect cells by scavenging excess copper.The prion protein (PrP) 1 is responsible for a novel class of infectious, neurodegenerative diseases collectively known as the transmissible spongiform encephalopathies (TSEs) (1-3). The TSEs include scrapie in sheep, mad cow disease (bovine spongiform encephalopathy, BSE), chronic wasting disease (CWD) in deer and elk, and Creutzfeldt-Jakob disease (CJD) in humans. The normal cellular form of the prion protein, referred to as PrP C , is found in a wide range of tissues of all mammalian and avian species. A misfolding event converts the protein to the infectious, β-sheet-rich scrapie form (PrP Sc ) responsible for the TSEs.PrP C is a GPI-anchored glycoprotein expressed in abundance on the surface of neurons, predominantly on presynaptic membranes (4,5). The mature form of PrP, consisting of residues 23-231 in hamster, has a globular C-terminal domain and a largely unstructured N-terminal domain (6) (Figure 1). The C-terminal domain (residues 125-231) contains three α helices, two of which are stabilized by a single interhelical disulfide bond, and a small antiparallel β sheet. The flexible N-terminal domain is glycine-rich and highly flexible. Within the flexible N-terminal region of PrP is the octarepeat domain, composed of four or five highly conserved, contiguous repeats of the eight-residue sequence PHGGGWGQ. The physiological function

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“…In some multimeric protein assemblies, these interactions may give rise to cooperative binding of substrates or ligands, which is believed to play a role in the regulation of their activity (Koshland & Hamadani, 2002).…”

Section: B Cooperativity and Cofactorsmentioning

confidence: 99%

Investigation of intact protein complexes by mass spectrometry

Heck

Heuvel

2004

Mass Spectrometry Reviews

559

554

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I. Introduction 00 II. Electrospray Ionization of Biomacromolecules 00 III. Mass Analyzers for Biomacromolecular Mass Spectrometry 00 A. Collisional Cooling and/or Focusing 00 B. Alternative Mass Analyzers 00 IV. Solution‐Phase Properties of Proteins Complexes 00 A. Protein–Protein Interactions 00 B. Cooperativity and Cofactors 00 C. Dynamics of Assembly 00 V. Gas‐Phase Properties of Protein Complexes 00 A. Tandem Mass Spectrometry 00 B. Charge Separation in Protein Dissociation 00 VI. Future Outlook 00 Acknowledgments 00 References 00 Mass spectrometry has grown in recent years to a well‐accepted and increasingly important complementary technique in structural biology. Especially electrospray ionization mass spectrometry is well suited for the detection of non‐covalent protein complexes and their interactions with DNA, RNA, ligands, and cofactors. Over the last decade, significant advances have been made in the ionization and mass analysis techniques, which makes the investigation of even larger and more heterogeneous intact assemblies feasible. These technological developments have paved the way to study intact non‐covalent protein–protein interactions, assembly and disassembly in real time, subunit exchange, cooperativity effects, and effects of cofactors, allowing us a better understanding of proteins in cellular processes. In this review, we describe some of the latest developments and several highlights. © 2004 Wiley Periodicals, Inc., Mass Spec Rev

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Proteomics and Models for Enzyme Cooperativity

Cited by 144 publications

References 90 publications

Insights into the Conformational Equilibria of Maltose-binding Protein by Analysis of High Affinity Mutants

Insights into the Conformational Equilibria of Maltose-binding Protein by Analysis of High Affinity Mutants

The Affinity of Copper Binding to the Prion Protein Octarepeat Domain: Evidence for Negative Cooperativity

Investigation of intact protein complexes by mass spectrometry

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