Infrared spectroscopy of proteins

This review discusses the application of infrared spectroscopy to the study of proteins. The focus is on the mid-infrared spectral region and the study of protein reactions by reaction-induced infrared difference spectroscopy.     

Orientational oscillations of the peptide groups of an α-helix

The low-energy orientational oscillations of the peptide groups of an -helix are considered and the value of the frequency is estimated to be in agreement with experiments. Approximate formulae are derived for the projection of a dipole moment on the helix axis and for the helix parameters. Within the framework of a three-chain model, the asymptotics of the soliton solution is obtained using a discrete approach.The analysis of -helix geometry exhibits two types of low-frequency oscillations of the -helix. The first one is connected with atom movements along the helix axis with the peptide groups twisting around the helix axis. Accordingly, it changes the hydrogen bond lengths between neighbouring peptide groups. In the second case, the slopes of the peptide groups to the helix axis oscillate without the helix parameters changing. Here, the energy of interactions between peptide-group dipoles is changed and, as a result, the oscillations have an optical nature. The frequency of the optical orientational oscillations is approximately 100 cm^-1.     

Voltage-dependent channel formation by rods of helical polypeptides

Summary The voltage-dependence of channel formation by alamethicin and its natural analogues can be described by a dipole flip-flop gating model, based on electric field-induced transbilayer orientational movements of single molecules. These field-induced changes in orientation result from the large permanent dipole moment of alamethicin, which adopts -helical conformation in hydrophobic medium. It was, therefore, supposed that the only structural requirement for voltage-dependent formation of alamethicin-type channels might be a rigid lipophilic helical segment of minimum length.In order to test this hypothesis we synthesized a family of lipophilic polypeptides—Boc-(Ala-Aib-Ala-Aib-Ala) _n -OMe,n=1–4—which adopt -helical conformation forn=2–4 and studied their interaction with planar lipid bilayers. Surprisingly, despite their large difference in chain length, all four polypeptides showed qualitatively similar behavior. At low field strength of the membrane electric field these polypeptides induce a significant, almost voltage-independent increase of the bilayer conductivity. At high field strength, however, a strongly voltage-dependent conductance increase occurs similar to that observed with alamethicin. It results from the opening of a multitude of ion translocating channels within the membrane phase.The steady-state voltage-dependent conductance depends on the 8^th–9^th power of polypeptide concentration and involves the transfer of 4–5 formal elementary charges. From the power dependences on polypeptide concentration and applied voltage of the time constants in voltage-jump current-relaxation experiments, it is concluded that channels could be formed from preexisting dodecamer aggregates by the simultaneous reorientation of six formal elementary charges. Channels exhibit large conductance values of several nS, which become larger towards shorter polypeptide chain length. A mean channel diameter of 19 Å is estimated corresponding roughly to the lumen diameter of a barrel comprised of 10 -helical staves. Similar to experiments with the N-terminal Boc-derivative of alamethicin we did not observe the burst sequence of nonintegral conductance steps typical of natural (N-terminal Ac-Aib)-alamethicin. Saturation in current/voltage curves as well as current inactivation in voltage-jump current-relaxation experiments are found. This may be understood by assuming that channels are generated as dodecamers but, while reaching the steady state, reduce their size to that of an octamer or nonamer. We conclude that the overall behavior of these synthetic polypeptides is very similar to that of alamethicin. They exhibit the same concentration and voltage-dependences but lack the stabilizing principle of resolved channel states characteristic of alamethicin.     

Power and work produced in different leg muscle groups when rising from a chair

The aim of this study was to determine the power output and work done by different muscle groups at the hip and knee joints during a rising movement, to be able to tell the degree of activation of the muscle groups and the relationship between concentric and eccentric work. Nine healthy male subjects rose from a chair with the seat at knee level. The moments of force about the hip and knee joints were calculated semidynamically. The power output (P) and work in the different muscle groups surrounding the joints was calculated as moment of force times joint angular velocity. Work was calculated as: work = f Pdt. The mean peak concentric power output was for the hip extensors 49.9 W, hip flexors 7.9 W and knee extensor 89.5 W. This power output corresponded to a net concentric work of 20.7 J, 1.0 J and 55.6 J, respectively. There was no concentric power output from the knee flexor muscles. Energy absorption through eccentric muscle action was produced by the hip extensors and hip flexors with a mean peak power output of 4.8 W and 7.4 W, respectively. It was concluded that during rising, the hip and knee muscles mainly worked concentrically and that the greatest power output and work were produced during concentric contraction of the knee and hip extensor muscles. There was however also a demand for eccentric work by the hip extensors as well as both concentric and eccentric work by the hip flexors. The knee flexor muscles were unloaded.     

Influence of the nature of the aromatic side-chain on the conductance of the channel of linear gramicidin: study of a series of 9,11,13,15-Tyr(O-protected) derivatives

This paper describes the single channel properties of a series of synthetic analogues of gramicidin A, where all four tryptophans are replaced either by tyrosine or by several O-protected (benzyl, methyl, ethyl or t-butyl) derivatives. It is shown that, although all analogues bear similar dipole moment on their side-chains, the conductance depends on the hydrophobicity of these protecting groups. An analysis of the conductance data suggests that the conductance is governed by the binding process and a possible explanation, based on conformational considerations, is proposed.Abbreviations GA X=tryptophane - GM X=phenylalanine - GN X=naphthylalanine - GQ8 X=8-quinolylalanine - GQ4 X=4-quinolylalanine - GT X=tyrosine - GTBzl X=O-benzyltyrosine - GTMe X=O-methyltyrosine - GTEt X=O-ethyltyrosine - GTBu X=O-t-butyltyrosine     

The effects of normalization method on antagonistic activity patterns during eccentric and concentric isokinetic knee extension and flexion

The purpose of this study was to compare different normalization methods of electromyographic (EMG) activity of antagonists during isokinetic eccentric and concentric knee movements. Twelve women performed three maximum knee extensions and flexions isometrically and at isokinetic concentric and eccentric angular velocities of 30 °·s⁻¹, 90 °·s⁻¹, 120 °·s⁻¹ and 150 °·s⁻¹. The EMG activity of the vastus lateralis, rectus femoris, vastus medialis and hamstrings was recorded. The antagonist integrated IEMG values were normalized relative to the EMG of the same muscle during an isometric maximal action (static method). The values were also expressed as a percentage of the EMG activity of the same muscle, at the same angle, angular velocity and muscle action (dynamic method) when the muscle was acting as an agonist. Three-way analysis of variance (ANOVA) designs indicated significantly greater IEMG normalized with the dynamic method compared to the EMG derived using the static method (P < 0.05). These differences were more evident at concentric angular velocities and at the first and last 20 ° of the movement. The present findings demonstrate that the method of normalization significantly influences the conclusions on antagonistic activity during isokinetic maximum voluntary efforts. The dynamic method of normalization is more appropriate because it considers the effects of muscle action, muscle length and angular velocity on antagonist IEMG.     

Motor unit recruitment strategy of knee antagonist muscles in a step-wise, increasing isometric contraction

The purpose of this study was to determine if differences exist between the control strategies of two antagonist thigh muscles during knee flexion and extension muscular coactivation. Surface myoelectric signal (MES) of the quadriceps (rectus femoris) and the hamstrings (semitendinosus) were obtained from both muscles while performing step-wise increasing contractions during flexion and extension with the knee at 1.57 rad of flexion (90 degrees). The median frequency of the power density spectrum, which is related to the average muscle fiber action potential conduction velocity and therefore to motor unit recruitment, was calculated from each MES. The results suggest that, in all the subjects tested, when the muscle acts as antagonist most motor units are recruited up to 50% of the maximal voluntary force, whereas when the muscle acts as antagonist motor units are recruited up to 40% of the maximal voluntary force. The force range past 40–50% of the maximal force is also characterized by differences between the agonist/antagonist.     

Comparison of the effects of hydrophobicity,amphiphilicity, and α-helicity on the activities of antimicrobial peptides

Multiple linear regression was used to quantify the dependence of the antimicrobial activity of 13 peptides upon three calculated or experimentally determined parameters: mean hydrophobicity, mean hydrophobic moment, and α-helix content. Mean hydrophobic moment is a measure of the amphiphilicity of peptides in an α-helical conformation. Antimicrobial activity was quantified as the reciprocal of the measured minimal inhibitory concentration (MIC) against Escherichia coli. One of the peptides was magainin 2, and the remainder were novel peptides designed for this study. The multiple linear regression results revealed that the amphiphilicity of the peptides was the most important factor governing anti-microbial activity compared to mean hydrophobicity orα-helix content. A better regression cf the data was obtained using In(1/MIC + constant) as the dependent variable than with either 1/MIC or In(1/MIC). These results should be useful in designing peptides with higher antimicrobial activity. © 1995 Wiley-Liss, Inc.     

Dielectric probe of the electric-field-sensitive peptide alamethicin

Dielectric constant and loss of alamethicin in solvents of various degrees of lipophilicity (namely mixtures of n-octanol and dioxane) have been measured at frequencies from 5 kHz to 50 MHz. By means of a conventional Cole-Cole approach, dielectric properties were evaluated to obtain information about the structural and dipolar properties of the peptide in view of its function as a voltage-dependent pore former in membranes. The results for a pure octanol solvent (together with an apparent molecular weight determined by ultracentrifugation) indicate the existence of primarily monomeric particles of quite elongated shape and of comparatively high dipole moment. Adding dioxane was found to yield considerable aggregation and a decrease of polarity.