Conformational peptides

Chapters focus on mass spectrometry, sequence and amino acid analysis, separations, protein folding and conformation, peptide and protein NMR, and peptide synthesis. In addition, the mutagenesis and protein design section has been expanded and a new section addresses analysis of protein interactions. 

Nachman, R. J., Zabrocki, J., Olczak, J., Williams, H. J., Moyna, G., Ian Scott, A., and Coast, G. M. (2002) cis-peptide bond mimetic tetrazole analogs of the insect kinins identify the active conformation. Peptides 23,709-716. 

The X-ray crystal structure database led us to believe that peptide bonds adopt either the cis or trans conformation in native proteins [22,128]. However, NMR spectroscopy [143], and in a few cases, crystal structure analysis [144], provide encouraging experimental evidence of conformational peptide bond polymorphism of folded proteins. Furthermore, conformational changes in response to ligand binding, crystallization conditions and point mutations at remote sites are frequent. Consequently, the three-dimensional protein structure database contains homologous proteins that have different native conformations for a critical prolyl bond [12]. 

Figure 1.17 Newman projections involving the N—bond and C —C(0) bond of a /S-strand to demonstrate the consequences of highly regular dihedral angles

CD has been used extensively to follow the equilibrium between helix and unordered conformation. Peptides undergoing helix-coil transitions typically exhibit an isodichroic point near 203 nm as the temperature, solvent, pH, or ionic strength is varied. The observation of an isodichroic... 

The catalytic subunit of cAPK contains two domains connected by a peptide linker. ATP binds in a deep cleft between the two domains. Presently, crystal structures showed cAPK in three different conformations, (1) in a closed conformation in the ternary complex with ATP or other tight-binding ligands and a peptide inhibitor PKI(5-24), (2) in an intermediate conformation in the binary complex with adenosine, and (3) in an open conformation in the binary complex of mammalian cAPK with PKI(5-24). Fig.l shows a superposition of the three protein kinase configurations to visualize the type of conformational movement. 

Fig. 2. Conformational free energy of closed, intermediate and open protein kinase conformations. cAPK indicates the unbound form of cAMP-dependent protein kinase, cAPKiATP the binary complex of cAPK with ATP, cAPKiPKP the binary complex of cAPK with the peptide inhibitor PKI(5-24), and cAPK PKI ATP the ternary complex of cAPK with ATP and PKI(5-24). Shown are averaged values for the three crystal structures lATP.pdb, ICDKA.pdb, and ICDKB.pdb. All values have been normalized with respect to the free energy of the closed conformations.

Conformational free energy simulations are being widely used in modeling of complex molecular systems [1]. Recent examples of applications include study of torsions in n-butane [2] and peptide sidechains [3, 4], as well as aggregation of methane [5] and a helix bundle protein in water [6]. Calculating free energy differences between molecular states is valuable because they are observable thermodynamic quantities, related to equilibrium constants and... 

The second application of the CFTI approach described here involves calculations of the free energy differences between conformers of the linear form of the opioid pentapeptide DPDPE in aqueous solution [9, 10]. DPDPE (Tyr-D-Pen-Gly-Phe-D-Pen, where D-Pen is the D isomer of /3,/3-dimethylcysteine) and other opioids are an interesting class of biologically active peptides which exhibit a strong correlation between conformation and affinity and selectivity for different receptors. The cyclic form of DPDPE contains a disulfide bond constraint, and is a highly specific S opioid [llj. Our simulations provide information on the cost of pre-organizing the linear peptide from its stable solution structure to a cyclic-like precursor for disulfide bond formation. Such... 

The two /3-turn structures, pc and Pe are the most stable among those considered. This is in accord with the unconstrained nanosecond simulations of linear DPDPE, which converged to these conformers [14]. Because the cyclic form is relatively rigid, it is assumed that the conformation it adopts in solution is the biologically active one, responsible for its high affinity and specificity towards the 5 opioid receptor. The relatively low population of the cyclic-like structure for the linear peptide thus agrees qualitatively with the... 

The Cyc conformer represents the structure adopted by the linear peptide prior to disulfide bond formation, while the two /3-turns are representative stable structures of linear DPDPE. The free energy differences of 4.0 kcal/mol between pc and Cyc, and 6.3 kcal/mol between pE and Cyc, reflect the cost of pre-organizing the linear peptide into a conformation conducive for disulfide bond formation. Such a conformational change is a pre-requisite for the chemical reaction of S-S bond formation to proceed. 

The free energy differences obtained from our constrained simulations refer to strictly specified states, defined by single points in the 14-dimensional dihedral space. Standard concepts of a molecular conformation include some region, or volume in that space, explored by thermal fluctuations around a transient equilibrium structure. To obtain the free energy differences between conformers of the unconstrained peptide, a correction for the thermodynamic state is needed. The volume of explored conformational space may be estimated from the covariance matrix of the coordinates of interest, = ((Ci [13, lOj. For each of the four selected conform-... 

Y. Wang and K. Kuczera. Exploration of conformational free energy surface of helical Ala and Aibn peptides. J. Phys. Chem. B, 101 5205-5213, 1997. 

Y. Wang and K. Kuczera. Conformational free energy surface of the linear DPDPE peptide Cost of pre-organization for disulfide bond formation. J. Am. Chem. Soc., submitted, 1997. 

The catalytic subunit then catalyzes the direct transfer of the 7-phosphate of ATP (visible as small beads at the end of ATP) to its peptide substrate. Catalysis takes place in the cleft between the two domains. Mutual orientation and position of these two lobes can be classified as either closed or open, for a review of the structures and function see e.g. [36]. The presented structure shows a closed conformation. Both the apoenzyme and the binary complex of the porcine C-subunit with di-iodinated inhibitor peptide represent the crystal structure in an open conformation [37] resulting from an overall rotation of the small lobe relative to the large lobe. 

Karlsson, R., Zheng, J., Zheng, N.-H., Taylor, S. S., Sowadski, J. M. Structure of the mamalian catalytic subunit of cAMP-dependent protein kinase and an inhibitor peptide displays an open conformation. Acta Cryst. D 49 (1993) 381-388. 

The Empirical Conformational Energy Program for Peptides, ECEPP [63, 64], is one of the first empirical interatomic potentials whose derivation is based both on gas-phase and X-ray crystal data [65], It was developed in 1975 and updated in 1983 and 1992. The actual distribution (dated May, 2000) can be downloaded without charge for academic use. 

D.R. Ripoll, H.A. Scheraga, ECEPP Empirical Conformational Energy Program for Peptides, in The Encyclopedia of Computational Chemistry, Vol. 2,... 

Molecular dynamics simulations are el ficient for searching the conformational space of medium-sized molecules and peptides. Different protocols can increase the elTicieiicy of the search and reduce the computer time needed to sample adequately the available conformations. 

High temperature searches of conformational space (see Quenched Dynamics" on page 78), can produce unwanted conformational changes, such as cis-tmnx peptide flips, ring inversions, and other changes that you cannot reverse easily by geometry optimization. You can use restraints to prevent these changes. 

The biologiccJ function of a protein or peptide is often intimately dependent upon the conformation(s) that the molecule can adopt. In contrast to most synthetic polymers where the individual molecules can adopt very different conformations, a protein usually exists in a single native state. These native states are found rmder conditions typically found in Uving cells (aqueous solvents near neutred pH at 20-40°C). Proteins can be unfolded (or denatured) using high-temperature, acidic or basic pH or certain non-aqueous solvents. However, this unfolding is often reversible cind so proteins can be folded back to their native structure in the laboratory. 

All of the conformational search methods that were described in Sections 9.2-9.7 have bee used at some stage to explore the conformational space of small pephdes. Here we wi describe some of the methods designed specifically for tackling the problem for peptide and proteins. 

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