High-throughput protein crystallography and drug discovery

Single crystal X-ray diffraction is the technique of choice for studying the interactions of small organic molecules with proteins by determining their three-dimensional structures; however the requirement for highly purified protein and lack of process automation have traditionally limited its use in this field. Despite these shortcomings, the use of crystal structures of therapeutically relevant drug targets in pharmaceutical research has increased significantly over the last decade. The application of structure-based drug design has resulted in several marketed drugs and is now an established discipline in most pharmaceutical companies. Furthermore, the recently published full genome sequences of Homo sapiens and a number of micro-organisms have provided a plethora of new potential drug targets that could be utilised in structure-based drug design programs. In order to take maximum advantage of this explosion of information, techniques have been developed to automate and speed up the various procedures required to obtain protein crystals of suitable quality, to collect and process the raw X-ray diffraction data into usable structural information, and to use three-dimensional protein structure as a basis for drug discovery and lead optimisation.This tutorial review covers the various technologies involved in the process pipeline for high-throughput protein crystallography as it is currently being applied to drug discovery. It is aimed at synthetic and computational chemists, as well as structural biologists, in both academia and industry, who are interested in structure-based drug design.     

Continuous-wave operation of a single-frequency optical parametric oscillator at 4-5 microm based on periodically poled LiNbO3

We present a cw, Nd:YAG-pumped singly resonant single-frequency narrow-linewidth high-power optical parametric oscillator with idler tuning from 3.7 to 4.7 microm. In this spectral range the absorption of the idler wave in the LiNbO3 crystal is significant, causing the oscillation threshold to increase with a subsequent decrease in output power from 1.2 W at 3.9 microm to 120 mW at 4.7 microm. The optical parametric oscillator's cavity was stabilized and mode-hop tuned with a rotatable solid etalon but with a subsequent reduction in idler power of as much as 50%. We demonstrated the usefulness for spectroscopy by recording the photoacoustic spectrum of a strong CO2 absorption, using a 24-GHz continuous idler scan.     

Rapid and sensitive trace gas detection with continuous wave Optical Parametric Oscillator-based Wavelength Modulation Spectroscopy

A fiber-amplified Distributed Bragg Reflector diode laser is used to pump a continuous wave, singly resonant Optical Parametric Oscillator (OPO). The output radiation covers the 3–4 μm with ability of rapid (100 THz/s) and broad mode-hop-free tuning (5 cm⁻¹). Wavelength Modulation Spectroscopy is combined with the OPO to take optimal advantage of the spectral scan speed. The sensitivity of the system was determined as 0.8 ppbv (parts-per-billion by volume) for ethane (C₂H₆) for the absorption peak at 2996.9 cm⁻¹ recorded in 1.3 seconds, corresponding to a noise equivalent absorption sensitivity (NEAS) of 1.2×10⁻⁹ cm⁻¹/Hz^1/2. A comparison between results using the 1^st, 2^nd and 4^th harmonic derivative signal from wavelength modulation was performed. The broad continuous tunability was demonstrated by covering 35 cm⁻¹ while recording absorption features of ethane, methane and water.     

Automatically tunable continuous-wave optical parametric oscillator for high-resolution spectroscopy and sensitive trace-gas detection

We present a high-power (2.75 W), broadly tunable (2.75–3.83 μm) continuous-wave optical parametric oscillator based on MgO-doped periodically poled lithium niobate. Automated tuning of the pump laser, etalon and crystal temperature results in a continuous wavelength coverage up to 450 cm^-1 per poling period at <5×10^-4 cm^-1 resolution. The versatility of the optical parametric oscillator as a coherent light source in trace-gas detection is demonstrated with photoacoustic and cavity ring-down spectroscopy. A 17-cm^-1-wide CO₂ spectrum at 2.8 μm and multi-component gas mixtures of methane, ethane and water in human breath were measured using photoacoustics. Methane (at 3.2 μm) and ethane (at 3.3 μm) were detected using cavity ring-down spectroscopy with detection limits of 0.16 and 0.07 parts per billion by volume, respectively. A recording of ¹²CH₄ and ¹³CH₄ isotopes of methane shows the ability to detect both species simultaneously at similar sensitivities. PACS 42.65.Yj; 42.72.Ai; 42.62.Fi     

Continuous‐wave optical parametric oscillator based infrared spectroscopy for sensitive molecular gas sensing

Over the past 10 years, with the advent of new crystals designs and a new generation of pump lasers, continuous‐wave (cw) optical parametric oscillators (OPOs) have developed into mature monochromatic light sources. Nowadays, cw OPOs can fulfill a wide variety of criteria for sensitive molecular gas sensing. It can access the mid‐infrared wavelength region, where many molecules have their fundamental rotational‐vibrational transitions, with high power. This high power combined with wide wavelength tuning and narrow linewidth creates excellent conditions for sensitive, high‐resolution spectroscopy. OPOs combined with robust methods, such as photoacoustic spectroscopy and cavity‐enhanced spectroscopy, are well suited for field measurements and remote‐sensing applications. The wide tunability of cw OPOs allows detection of larger molecules with broad absorption band structures, and its fast scanning capabilities allow rapid detection of trace gases, the latter is a demand for life‐science applications. After a short introduction about the physical principle of cw OPOs, with its most recent physical developments, this review focuses on sensitive molecular gas sensing with a variety of spectroscopic applications in atmospheric and life sciences.     

Thermal effects in singly resonant continuous-wave optical parametric oscillators

Stability and tuning characteristics of continuous-wave optical parametric oscillators (CW OPOs) are affected by various thermal effects arising from optical absorption in nonlinear crystals. In this paper, we present an experimental study of such effects in a singly resonant CW OPO. The OPO operates in the 3-μm mid-infrared region and it is based on a MgO-doped periodically poled lithium niobate crystal. We focus our study on two thermally induced phenomena that have been recently reported to exist in singly resonant CW OPOs: optical bi-stability and thermal self-locking. Thermal self-locking effect, which is known to alter the stability and tuning properties of doubly and triply resonant CW OPOs, is shown to be also of importance in singly resonant OPOs. We report the stability and tuning characteristics of a thermally loaded OPO and discuss a simple temperature-tuning method that can be used to scan the OPO idler frequency continuously over several THz.     

CO-laser-based photoacoustic trace-gas detection: applications in postharvest physiology

2 measurements using polarographic oxygen sensors. Received: 13 March 1998/Revised version: 13 May 1998     

Photoacoustic spectroscopy using quantum-cascade lasers

Photoacoustic spectra of ammonia and water vapor were recorded by use of a continuous-wave quantum-cascade distributed-feedback (QC-DFB) laser at 8.5 mum with a 16-mW power output. The gases were flowed through a cell that was resonant at 1.6 kHz, and the QC-DFB source was temperature tuned over 35 nm for generation of spectra or was temperature stabilized on an absorption feature peak to permit real-time concentration measurements. A detection limit of 100 parts in 10(9) by volume ammonia at standard temperature and pressure was obtained for a 1-Hz bandwidth in a measurement time of 10 min.