Local Conformational Differences Research Articles

EPR and fluorescence depolarization studies on bovine cardiac myosin.

To test for possible differences in local conformation and S1 flexibility, bovine cardiac and rabbit skeletal myosins were labeled with a fluorophore (1,5-IAEDANS) and a spin label having iodoacetamide reactivity. The marked activation of the Ca2+-ATPase (6- to 8-fold) and inhibition of the K+ (EDTA)-ATPase (80-90%) by both labels indicated specific labeling of the fast-reacting thiols (SH1) of both myosins. Fluorescence depolarization studies of 1,5-IAEDANS-labeled cardiac myosin indicated that, like skeletal myosin, the SI moieties of cardiac myosin exhibit considerable segmental flexibility with respect to the rod portion of the molecule. This indicates that segmental flexibility may be a property of all myosins. Cardiac and skeletal myosins immobilized spin labels to approximately the same extent, indicating a similarity in steric restraints around the SH1 thiol of the two myosins. The magnitude of the changes in spin label mobility accompanying binding of MgADP and hydrolysis of MgATP was reduced in cardiac myosin relative to skeletal myosin. This suggests that the lower catalytic center activity of cardiac myosin is associated with more restricted conformational changes accompanying formation of M.ADP and M.ADP.Pi. From measurements of spin label mobility, the affinity of cardiac and skeletal myosin for ADP were similar: Kd (ADP) = 7 microM, n = 1.6. The EPR spectrum of spin labels attached to cardiac and skeletal myosin showed similar saturation effects upon actin binding indicating immobilization of myosin heads occurs with both proteins.

The crystal structure of monoclinic ribonuclease-S at six Ångstroms resolution

An independent structure analysis has been made of ribonuclease-S crystallized in a monoclinic space group C2 at 6 Å resolution. The conformations of the two crystallographically independent molecules (molecule ZA and ZB) were compared with that of a chemically identical molecule (molecule Y) crystallized in a trigonal space group P3 121, the structure of which has been solved to 2.0 Å resolution by Wyckoff et al. (1970). The N-terminal tail of the S-protein of molecule ZA assumes a unique conformation somewhat resembling that of ribonuclease-A, while the corresponding part of molecule ZB assumes about a similar conformation to that of molecule Y. Apart from the solvated terminal region, the overall arrangements of various features of the three structures are very similar, although possibilities of local conformational differences are not considered at this stage of the analysis. The environments of the three molecules in the crystal lattice are compared in detail. Two of the four molecules in the primitive cell are related to each other by a crystallographic 2-fold axis very similar to a 2-fold relationship found in the Y-form. All other relationships are quite different.

Conformational studies of poly(methacrylic Acid). II. Laser-excited Raman studies of the conformational transition in aqueous solution

Abstract The conformational transitions of atactic, syndiotactic, and isotactic poly-(methacrylic acid) in aqueous solution as a function of sodium hydroxide neutralization were investigated with laser-excited Raman spectroscopy. It was concluded that the conformational transition of the syndiotactic and atactic polymer occurs gradually over the range of approximately 20 to 50% neutralization and cannot be considered to be cooperative. However, the conformational transition of the isotactic polymer occurs over a much narrower neutralization range and may be cooperative. Raman spectra of the various stereoregular forms indicate a lack of local helical order. The Raman spectra of the polyacids also indicate no large differences in local conformation for the different stereoregular forms, a result that is in agreement with conformational energy analysis of isolated isotactic and syndiotactic chains. However, Raman spectra of the stereoregular forms of poly(sodium methacrylate) are markedly different, indicat...