Crossing the Border: From Keto- to Imine Reduction in Short-Chain Dehydrogenases/Reductases

The family of NAD(P)H-dependent short-chain dehydrogenases/reductases (SDRs) comprises numerous biocatalysts capable of C=O or C=C reduction. The highly homologous noroxomaritidine reductase (NR) from Narcissus sp. aff. pseudonarcissus and Zt_SDR from Zephyranthes treatiae, however, are SDRs with an extended imine substrate scope. Comparison with a similar SDR from Asparagus officinalis (Ao_SDR) exhibiting keto-reducing activity, yet negligible imine-reducing capability, and mining the Short-Chain Dehydrogenase/Reductase Engineering Database indicated that NR and Zt_SDR possess a unique active-site composition among SDRs. Adapting the active site of Ao_SDR accordingly improved its imine-reducing capability. By applying the same strategy, an unrelated SDR from Methylobacterium sp. 77 (M77_SDR) with distinct keto-reducing activity was engineered into a promiscuous enzyme with imine-reducing activity, thereby confirming that the ability to reduce imines can be rationally introduced into members of the “classical” SDR enzyme family. Thus, members of the SDR family could be a promising starting point for protein approaches to generate new imine-reducing enzymes.     

Ultrasonic technique for measuring porosity of plasma-sprayed alumina coatings

Porosity is an important factor in plasma-sprayed coatings, especially ceramic coatings. Excessive poros-ity can adversely affect the performance of the coated component in various ways. An ultrasonic nonde-structive measurement technique has been developed to measure porosity in plasma-sprayed alumina coatings. The technique is generic and can be extended to other ceramic coating systems. To test the tech-nique, freestanding alumina coatings with varying levels of porosity were fabricated via plasma spray. Samples with varying porosity, obtained through innovative fabrication techniques, were used to gener-ate a calibration curve. The ultrasonic velocity in the low-frequency range was found to be dependent on the density of freestanding coatings (measured via Archimedian techniques). This dependence is the basis of the development of a technique to measure the density of coatings.     

Fatigue lifetime prediction with the aid of SAW NDE

The present studies concentrated on predicting the remaining fatigue life for single fatigue cracks in the Paris regime of macrocrack propagation. Acoustic surface waves were used to interrogate the crack during cyclic fatigue. The inversion of the obtained scattering data provided crack depth and crack length as a function of the number of cycles applied in tension-tension fatigue. Auxiliary experiments were conducted to study the acoustic response of the crack to tensile and compressive loads, thought to open and close the crack. The technique may allow for new insights into the physics of the crack closure effect.     

Design of acoustic beam aperture modifier using gradient-index phononic crystals

This article reports the design concept of a novel acoustic beam aperture modifier using butt-jointed gradient-index phononic crystals (GRIN PCs) consisting of steel cylinders embedded in a homogeneous epoxy background. By gradually tuning the period of a GRIN PC, the propagating direction of acoustic waves can be continuously bent to follow a sinusoidal trajectory in the structure. The aperture of an acoustic beam can therefore be shrunk or expanded through change of the gradient refractive index profiles of the butt-jointed GRIN PCs. Our computational results elucidate the effectiveness of the proposed acoustic beam aperture modifier. Such an acoustic device can be fabricated through a simple process and will be valuable in applications, such as biomedical imaging and surgery, nondestructive evaluation, communication, and acoustic absorbers.     

Reduction of losses in low-frequency (2 MHz) surface-wave delay line by outgassing

Output signal enhancements by as much as 8 dB were achieved for 2.25 MHz surface-wave delay line (55 ?s delay) made of high-density p.z.t. by outgassing treatments in vacuum.     

High sensitivity fiber optic angular displacement sensor and its application for detection of ultrasound

In this paper, we report on the development of an intensity-modulated fiber-optic sensor for angular displacement measurement. This sensor was designed to present high sensitivity, linear response, and wide bandwidth and, furthermore, to be simple and low cost. The sensor comprises two optical fibers, a positive lens, a reflective surface, an optical source, and a photodetector. A mathematical model was developed to determine and simulate the static characteristic curve of the sensor and to compare different sensor configurations regarding the core radii of the optical fibers. The simulation results showed that the sensor configurations tested are highly sensitive to small angle variation (in the range of microradians) with nonlinearity less than or equal to 1%. The normalized sensitivity ranges from (0.25×V(max)) to (2.40×V(max)) mV/μrad (where V(max) is the peak voltage of the static characteristic curve), and the linear range is from 194 to 1840 μrad. The unnormalized sensitivity for a reflective surface with reflectivity of 100% was measured as 7.7 mV/μrad. The simulations were compared with experimental results to validate the mathematical model and to define the most suitable configuration for ultrasonic detection. The sensor was tested on the characterization of a piezoelectric transducer and as part of a laser ultrasonics setup. The velocities of the longitudinal, shear, and surface waves were measured on aluminum samples as 6.43, 3.17, and 2.96 mm/μs, respectively, with an error smaller than 1.3%. The sensor, an alternative to piezoelectric or interferometric detectors, proved to be suitable for detection of ultrasonic waves and to perform time-of-flight measurements and nondestructive inspection.     

Acoustic imaging of thick biological tissue

Up to now, biomedical imaging with ultrasound for observing a cellular tissue structure has been limited to very thinly sliced tissue at very high ultrasonic frequencies, i.e., 1 GHz. In this paper, we present the results of a systematic study to use a 150 to 200 MHz frequency range for thickly sliced biological tissue. A mechanical scanning reflection acoustic microscope (SAM) was used for obtaining horizontal crosssectional images (C-scans) showing cellular structures. In the study, sectioned specimens of human breast cancer and tissues from the small intestine were prepared and examined. Some accessories for biomedical application were integrated into our SAM (Sonix HS-1000 and Olympus UH-3), which operated in pulse-wave and tone-burst wave modes, respectively. We found that the frequency 100 to 200 MHz provides optimal balance between resolution and penetration depth for examining the thickly sliced specimens. The images obtained with the lens focused at different depths revealed cellular structures whose morphology was very similar to that seen in the thinly sectioned specimens with optical and scanning acoustic microscopy. The SAM operation in the pulse-echo mode permits the imaging of tissue structure at the surface, and it also opens up the potential for attenuation imaging representing reflection from the substrate behind the thick specimen. We present such images of breast cancer proving the method?s applicability to overall tumor detection. SAM with a high-frequency tone-burst ultrasonic wave reveals details of tissue structure, and both methods may serve as additional diagnostic tools in a hospital environment.     

Tuning and Switching Enantioselectivity of Asymmetric Carboligation in an Enzyme through Mutational Analysis of a Single Hot Spot

Enantioselective bond making and breaking is a hallmark of enzyme action, yet switching the enantioselectivity of the reaction is a difficult undertaking, and typically requires extensive screening of mutant libraries and multiple mutations. Here, we demonstrate that mutational diversification of a single catalytic hot spot in the enzyme pyruvate decarboxylase gives access to both enantiomers of acyloins acetoin and phenylacetylcarbinol, important pharmaceutical precursors, in the case of acetoin even starting from the unselective wild‐type protein. Protein crystallography was used to rationalize these findings and to propose a mechanistic model of how enantioselectivity is controlled. In a broader context, our studies highlight the efficiency of mechanism‐inspired and structure‐guided rational protein design for enhancing and switching enantioselectivity of enzymatic reactions, by systematically exploring the biocatalytic potential of a single hot spot.     

Laser-based ultrasonics on Gr/epoxy composite

Critical issues are examined in the application of laser generation and detection of ultrasound to the inspection of large area air-frame composites. Among these issues are surface roughness, signal-to-noise ratio, insensitivity to the path length between the part and detector, and wide band vs. narrow-band generation. Demonstrated is the feasibility of transmission C-scans of 150 ply Gr/epoxy panels containing simulated delaminations. Waveforms having a signal-to-noise ratio of greater than 40 dB in 15-mm thick Gr/epoxy were obtained with generating laser powers well below the ablation limit for the graphite epoxy. Detection was shot-noise limited with a detector noise figure of about 5–10 dB.