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The resistance of Gram-negative bacteria to β-lactam antibiotics stems mainly from β-lactamase proteins that hydrolytically deactivate the β-lactams. Of particular concern are the β-lactamases that can deactivate a class of β-lactams known as carbapenems. Carbapenems are among the few anti-infectives that can treat multi-drug resistant bacterial infections. Revealing the mechanisms of their deactivation by β-lactamases is a necessary step for preserving their therapeutic value. Here, we present NMR investigations of OXA-24/40, a carbapenem-hydrolyzing Class D β-lactamase (CHDL) expressed in the gram-negative pathogen, Acinetobacter baumannii. Using rapid data acquisition methods, we were able to study the “real-time” deactivation of the carbapenem known as doripenem by OXA-24/40. Our results indicate that OXA-24/40 has two deactivation mechanisms: canonical hydrolytic cleavage, and a distinct mechanism that produces a β-lactone product that has weak affinity for the OXA-24/40 active site. The mechanisms issue from distinct active site environments poised either for hydrolysis or β-lactone formation. Mutagenesis reveals that R261, a conserved active site arginine, stabilizes the active site environment enabling β-lactone formation. Our results have implications not only for OXA-24/40, but the larger family of CHDLs now challenging clinical settings on a global scale.

Gram-negative bacteria resist β-lactam antibiotics primarily by deploying β-lactamase proteins that hydrolytically destroy the antibiotics. In clinical settings, these bacteria are producing variant β-lactamases with "gain-of-activity" mutations that inactivate a broader range of β-lactams. Learning how these mutations broaden substrate activity is important for coping with β-lactam resistance. Here, we investigate a gain of activity mutation in OXA-24/40, a carbapenem-hydrolyzing class D β-lactamase (CHDL) in Acinetobacter baumannii. OXA-24/40 was originally active against penicillin and carbapenem classes of β-lactams, but a clinical variant of OXA-24/40, the single-site substitution mutant P227S, has emerged with expanded activity that now includes advanced cephalosporins and the monobactam aztreonam. Using solution-state nuclear magnetic resonance (NMR) spectroscopy, we have compared the site-specific backbone dynamics of wild-type OXA-24/40 and the P227S variant. P227S changes local backbone flexibility in segments that are important for both binding and hydrolysis of carbapenem and cephalosporin substrates. Our results suggest that mutation-induced changes in sequence-specific dynamics can expand substrate activity and thus highlight the role of protein conformational dynamics in antibiotic resistance. To the best of our knowledge, this is the first NMR study of CHDL conformational dynamics and its impact on the expansion of β-lactam antibiotic resistance.

The transmembrane antibiotic sensor/signal transducer protein BlaR1 is part of a cohort of proteins that confer β-lactam antibiotic resistance in methicillin-resistant Staphylococcus aureus (MRSA) [Fisher, J. F., Meroueh, S. O., and Mobashery, S. (2005) Chem. Rev. 105, 395-424; Llarrull, L. I., Fisher, J. F., and Mobashery, S. (2009) Antimicrob. Agents Chemother. 53, 4051-4063; Llarrull, L. I., Toth, M., Champion, M. M., and Mobashery, S. (2011) J. Biol. Chem. 286, 38148-38158]. Specifically, BlaR1 regulates the inducible expression of β-lactamases that hydrolytically destroy β-lactam antibiotics. The resistance phenotype starts with β-lactam antibiotic acylation of the BlaR1 extracellular domain (BlaRS). The acylation activates the cytoplasmic protease domain through an obscure signal transduction mechanism. Here, we compare protein dynamics of apo versus antibiotic-acylated BlaRS using nuclear magnetic resonance. Our analyses reveal inter-residue interactions that relay acylation-induced perturbations within the antibiotic-binding site to the transmembrane helix regions near the membrane surface. These are the first insights into the process of signal transduction by BlaR1.

A version of a scheme of speckle interferometry is considered, which is intended for the analysis of the stationary distribution pattern of steep refractive-index gradients in optically transparent media. The application of this approach to problems of hydrodynamics and gas dynamics allows one to determine the structure of distribution of optical inhomogeneities in liquid and gaseous flows or media. The method is based on the recording of patterns of two speckle structures obtained in the plane of image formation of the object by an optical system when the object is illuminated by radiation having an auxiliary speckle structure. The patterns correspond to the two positions of an object shifted somewhat in the direction of the optical axis of the system. Contrary to the common methods of speckle interferometry, only the state of the object being investigated for two different conditions of illumination by the probe radiation is used for the detection in this case. The difficulties arising in the measurements of the objects with steep refractive-index gradients by the known methods of speckle interferometry are accordingly practically eliminated in the scheme under consideration. In comparison with the schlieren phase methods, this method offers higher possibilities in the variation of sensitivity.

A version of a scheme of speckle interferometry is considered, which is intended for the analysis of the distribution of refractive-index gradients in optically transparent media. The application of this approach to problems of hydrodynamics and gas dynamics makes it possible to determine the structure of the distribution of optical inhomogeneities in liquid and gaseous flows. The method is based on the recording of patterns of two speckle structures obtained in the plane of spatial frequencies by using illumination with an auxiliary speckle structure. One speckle structure is formed in the absence of an object, and the other one is formed by transilluminating an object. In comparison with other known phase methods, the speckle-interferometry methods make it possible to change the measurement sensitivity in a wide range and substantially simplify the interpretation of the results obtained.

The correct interpretation of light scattering data requires knowledge of the extent to which multiple scattering contributes to the measured signal. Generally, this information cannot easily be obtained in conventional light scattering experiments. We demonstrate the difficulties which arise with this problem by experiments with scatterers where the differential scattering cross section does not exhibit scattering minima. We investigate the influence of multiple scattering by performing combined static and dynamic light scattering experiments using the 3D cross correlation technique. We discuss that, despite the increase of the intensity ratio of multiply to singly scattered light with increasing scattering angle, the dynamic properties are more strongly affected for small scattering angles.

An optical method to measure velocity gradients was developed further to measure temporal correlation functions of the components of the velocity gradient tensor. For a turbulent channel flow (Reynolds number 3850) all non-vanishing correlation functions were obtained. Some quantities of first-order statistics such as for instance the turbulent energy dissipation and variances and covariances of the velocity gradient as well as quantities of second-order statistics such as vorticity correlation functions are presented.

Space- and time-dependent stochastic processes can exhibit correlated spatio-temporal fluctuations, which in special cases can be interpreted as the velocity of the process. We show how for a certain class of such processes - time dependent two-dimensional stochastic patterns - this velocity can be determined experimentally. Since the experimental method is based on differential properties of the pattern, we have calculated the joint distribution function of these quantities for a common type of circular Gaussian process.

We measured the distance‐dependent coupling of transverse boundary waves in a system of layers of two immiscible liquids by laser light scattering. The results were explained on the basis of a theoretical treatment of the dynamics of this system. The deviations of the results for very small distances of the two interfaces could be accounted for by the assumption of a small surface viscoelasticity, which is about one order of magnitude smaller than those measured so far with light‐scattering techniques. The technique used in this paper might therefore be well suited to study molecular interactions and relaxation processes at liquid boundaries, which are responsible for this viscoelasticity.

A light scattering technique is described which allows the non-invasive measurement of instantaneous velocity gradient components in fluid flow. Statistical and systematic errors are investigated both experimentally and theoretically. The method represents a basis for the study of temporal evolution of velocity gradients in fluid flow.

The motion of the speckle pattern in the far field limit is uniquely determined by the velocity gradient of the light scattering medium. This is the basic physical effect of a non-invasive method that allows the measurement of velocity gradients with high spatial and temporal resolution down to 75 μm and 1 ms, respectively. We present three different measurement schemes for the analysis of the scattered light. The experimental results refer to mean gradients obtained from measurements on turbulent channel flows.

A laser light scattering technique is described which allows the measurement of velocity gradients and their temporal correlation functions in fluid flow. This non-invasive and gauge-free technique was applied to a turbulent channel flow. Results of the measurements, including the turbulent energy dissipation and the enstrophy are presented.

Random spatio-temporal intensity patterns occur especially in the field of laser light scattering. In many cases the spatial motion of the intensity field of the scattered light contains information on the scatterer (speckle anemometry). We propose a new technique for measuring this velocity, which is especially suited for fast changing patterns (fast speckle boiling), and show experimental results.

A method of measuring velocity gradients on the basis of a photon correlation technique has been developed. Depending on the scattering geometry, particular rows of the velocity gradient tensor Gamma can be determined for laminar flow without any knowledge of the flow symmetry. The method is also applicable to turbulent flow. With the assumption of a normal distribution of the tensor elements of one row of Gamma , the measured correlation functions for a tube flow can be fitted satisfactorily. Limits of the applicability of the method are discussed.

An interferometric method in connection with a correlation technique for measuring the velocity of moving parts of plants is described. Velocities down to 5-0 mm d_1 can be measured reliably in a minute. Values of stem elongation rates of etiolated seedlings of a variety of species obtained by this method are very similar to those obtained by the application of linear displacement transducers. Since the optical method requires no mechanical coupling to the plant, velocities of sensitive plant organs can be determined, as shown by measurements of leaf motion.

If a laser beam passes a liquid sample a small fraction of the incident power is always absorbed by the medium. This gives rise to a temperature profile causing gradients of the density and of the index or refraction, and it changes the optical properties of the sample. The theory of this effect (thermal lens) is generalized for thick samples and focussed laser beams. An analytical solution for the beam properties is given as a function of the position in the sample. The theory is applied to light scattering experiments and analytical expressions are given for the time dependence of the intensity of the scattered light, depending on the power of the incident laser beam, experimental layout, absorbance and the thermal conductivity of the sample. The stationary solution for the intensity of the scattered light contains a contribution which is quadratic in the intensity of the incident laser beam. The theory is verified experimentally. Consequence for experiments where the intensity of the incident laser beam varies with time or where the sample itself shows time-dependent properties are discussed.

Measurements of the Rayleigh-Brillouin spectra of the binary system nitroethane-isooctane have been carried out as function of temperature and concentration. The purpose of this investigation was to study the transition of one type of phase transformation to an other. Taking the temperature as variable, the phase transformation is of second order for one specific concentration – the critical concentration, whereas for other concentrations it is of first order. Samples deviating up to 20% from the critical composition have been investigated in the temperature range 10 to 10−3 K above the transition temperature in the homogeneous region. The temperature dependence of the Rayleigh-Brillouin intensity ratio Ir/2Ib can be described by the same critical exponent γ = 1.24 for all concentrations provided the critical temperature in the scaling function is replaced by another reference temperature, interpreted as pseudo-spinodal temperature T*s The concentration dependence of T*s is found to be |X – Xc| = A · Tβ*s, where ts is the reduced temperature ts = 1 –T*s/Tc. The value of the exponent is found to be β* = 0.5, which is the classical exponent of the spinodal curve.

The influence of an external magnetic field on the normal incidence reflectivity of CuCl single crystals has been measured in the Voigt and Faraday configurations. The results are discussed in terms of the dielectric function calculated from the reflectivity spectrum by a Kramers-Kronig transformation. In the Voigt configuration two new exciton bands can be observed, which are attributed to a triplet and a longitudinal exciton. Their energetic positions are used to determine the parameters of electron-hole exchange interaction in the 1s excitons, and the spin-orbit splitting parameter of the upper valence band. The excitonic g-values are determined from the Faraday configuration measurements and can be interpreted on the basis of the tight binding model.

Low temperature crystallographic phase transitions in DyVO4 and TbVO are attributed to a co-operative Jahn-Teller effect. A preliminary report of the theory is given, based on a harmonic oscillator Hamiltonian for the crystal and an electron lattice coupling which is small compared to typical phonon zero point energies. An Ising Hamiltonian is shown to describe the co-operative ordering.

A Hamiltonian for the co-operative Jahn-Teller transition in DyVO4 is derived with both optic mode and strain couplings. A molecular field theory for this Hamiltonian predicts a second order phase transition and a dependence of the ground state splitting on stress. Contributions of phonon modes and strain can be distinguished by a comparison of the elastic constants and the parameters derived from the stress dependence of the ground state splitting. A correction to these molecular field results is derived by accounting for fluctuations. The effects of a magnetic field are investigated in the molecular field approximation; it is found that if the field is applied in the basal plane simultaneously with the stress at low temperatures, different distortions can be stabilized. The form of the first order phase boundary for these distortions is derived and an analogy is drawn between this ‘pseudospin flop’ transition and antiferromagnetic spin flop.

The magnetic circular dichroism of the K-band in the alkali halides RbCl, RbBr, RbI, KCl, KBr, and KI has been measured at temperatures between 4.2 and 77 °K. The magnetic field up to 50 kG was produced by a superconducting solenoid. We interpret the experimental results using the method of moments. The spin-orbit splitting and the orbital g-factors of the excited states due to the K-band are determined. Furthermore we obtain matrix elements describing the mixture of the excited states giving rise to F-and K-bands. In accordance with calculations of Smith [15] the measurements confirm the Mott-Gurney model, which attributes the K-band mainly to 1s-3p transitions. Der magnetische zirkulare Dichroismus der K-Bande wurde in den Alkalihalogeniden RbCl, RbBr, RbJ, KCl, KBr und KJ bei Temperaturen von 4.2 bis 77 °K gemessen. Als Magnet diente eine Supraleitungsspule, mit der Magnetfelder bis zu 50 kG erzeugt werden konnen. Die Auswertung der Messungen geschieht nach der Methode der Momente. Man erhalt Spin-Bahn-Aufspaltungen und orbitale g-Faktoren der angeregten Zustande des K-Uberganges. Auserdem werden Matrixelemente bestimmt, die die Mischung der angeregten Zustande des K- und F-Uberganges beschreiben. In Ubereinstimmung mit Rechnungen von Smith [15] bestatigen die Messungen das Mott-Gurney-Modell, das die K-Bande im wesentlichen 1s-3p-Ubergangen zuordnet.

Optical absorption measurements on TbVO 4 indicate that at 33°K there is a second order crystallographic phase change. Although no magnetic ordering occurs, the distortion direction can be controlled by a magnetic field. This behaviour is compared to DyVO 4 , in which a similar distortion occurs along different crystallographic directions.

Non-homogeneous velocity distributions of light scattering particles generally result in a non-homogeneous motion of the speckle pattern. This has been shown theoretically and experimentally by considering the space-time intensity correlation function of light being scattered by particles within a shear flow.

T1I-layers with simple cubic and f.c.c. structure have been prepared by evaporation of CsI and RbI. The optical absorption spectra of these layers have been measured for the first time and are shown to be very similar to those of simple cubic and f.c.c. T1C1 and T1Br, respectively. Relativistic band-structure calculations explain the optical spectra as well as these similarities.

The application of dynamic light scattering to flow measurements has proven a valuable tool in order to determine cartesian velocity components of a flow seeded with light scattering particles. Main advantages compared to other methods are that, on the one hand, there is no influence of the measuring device on the flow, and on the other hand there is no gauging necessary because only geometrical factors enter the final result.

Transverse magnetoreflection measurements give evidence for a longitudinal and a triplet state near the first exciton band in CuCl.

Optical absorption spectroscopy is used to investigate the low lying energy levels of DyVO 4 under uniaxial stress and magnetic fields at temperatures above and below the co-operative Jahn-Teller transition. The results agree with the molecular field theory of the previous paper using the parameter σ 0 = 64 ± 4 kg/mm 2 (which represents the stress of e aa − e bb type required to split the ground state energy levels by 27 cm −1 at high temperatures). The behaviour of dilute Dy 3+ in YVO 4 is studied and predicted from the theory. Deviations from the molecular field theory are found and discussed. Comparison of the results with known elastic constants of DyVO 4 indicates that strain coupling dominates the optic mode coupling.

Circular dichroism measurements were used to determine the spin orbit splitting Δ and the orbital g -factor g orb of the excited state belonging to the K absorption in RbCl.