Biomedical Spectroscopy and Imaging - Volume 7, issue 1-2
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Biomedical Spectroscopy and Imaging (BSI) is a multidisciplinary journal devoted to the timely publication of basic and applied research that uses spectroscopic and imaging techniques in different areas of life science including biology, biochemistry, biotechnology, bionanotechnology, environmental science, food science, pharmaceutical science, physiology and medicine. Scientists are encouraged to submit their work for publication in the form of original articles, brief communications, rapid communications, reviews and mini-reviews.
The journal is dedicated to providing a single forum for experts in spectroscopy and imaging as applied to biomedical problems, and also for life scientists who use these powerful methods for advancing their research work. BSI aims to promote communication, understanding and synergy across the diverse disciplines that rely on spectroscopy and imaging. It also encourages the submission of articles describing development of new devices and technologies, based on spectroscopy and imaging methods, for application in diverse areas including medicine, biomedical science, biomaterials science, environmental science, pharmaceutical science, proteomics, genomics, metabolomics, microbiology, biotechnology, genetic engineering, nanotechnology, etc.
Abstract: This article reviews work presented at the ESCBM 2017 on using two-dimensional infrared (2D IR) spectroscopy to probe ion binding configurations in the potassium ion channel KcsA. We discuss two studies in which we use 2D IR spectroscopy in conjunction with protein semisynthesis and molecular dynamics (MD) simulations to test against two competing models of ion permeation and to investigate how changes in the conformation of the intracellular gate affect the structure and ion affinities of the selectivity filter. In our initial study, the 2D IR spectrum of an isotopically labeled KcsA filter reveals two spectral features that correspond to…different structures and ion binding configurations. MD simulations help us link the experiments to atomistic structures, allowing us to determine the prevailing mechanism of ion conduction. In a follow-up study, we probe how ion occupancies in the filter change with the opening and closing of the intracellular gate. We experimentally modulate the conformation of KcsA using different sample conditions, and are able to show that the ion occupancies in the filter change with the state of the gate, revealing complexities in the conformational landscape of the potassium ion channel. Our work shows the potential of these techniques in addressing the fundamental biophysical questions in membrane protein structure and function.
Keywords: 2D IR spectroscopy, potassium ion channels, ion permeation, protein semisynthesis
Abstract: Solid supported lipid bilayers (SSLB) play an important role as biomimetic membranes to study protein-membrane interactions. We investigated the orientation of lipids in SSLBs at different temperatures and over time. Especially the stability of the lipid bilayer and structural changes upon lipid phase transition were analyzed by polarized ATR-FTIR spectroscopy and with SSLBs of different lipid compositions. The integrity of a lipid bilayer consisting of POPC or a 1:1 mixture of POPC and POPG is conserved over a wide temperature range and over several hours. Furthermore, we were able to monitor changes in the orientation of the lipid alkyl chains…upon lipid phase transition for DMPC and DSPC. This study shows that the combination of solid supported lipid bilayers and polarized ATR-FTIR spectroscopy is very powerful to characterize lipid membranes under different environmental conditions. The sensitivity of this technique will be exploited in future studies to analyze the effect of protein-membrane interaction on lipid orientation.
Abstract: Attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy is a surface-sensitive and label-free technique, which is applied to obtain dynamic structural information of biomolecules. The study of proteins by ATR-FTIR spectroscopy can be impeded by their tendency to adsorb to solid surfaces. Furthermore, the adsorption process of proteins is often accompanied with conformational changes, which can interfere with the intended structural analysis. We efficiently modified a silicon ATR crystal surface with polyethylene glycol and thereby create a protein-repellent surface. To achieve a high sensitivity, which enables the study of small conformational changes of biomolecules, we combine surface passivation with specific immobilization.…This is accomplished via the biotin-streptavidin interaction, which is one of the strongest known non-covalent protein-ligand interactions. As a proof of concept we present the specific immobilization of DNA. The modified surface is stable against elevated temperatures and 8 M urea and can therefore be used to study a wide range of biochemical systems and reactions. The surface chemistry is simple and performed under mild conditions, which leads to a high applicability of the presented approach.
Keywords: ATR-FTIR spectroscopy, biomolecules, protein adsorption, surface passivation, specific immobilization
Abstract: Background: High emission powers of external cavity-quantum cascade laser (EC-QCL) light sources allow to employ significantly larger path lengths for infrared (IR) transmission measurements compared to conventional Fourier-transform infrared (FTIR) measurements employing thermal emitters. Objective: An EC-QCL based IR transmission setup is presented as a viable alternative for analysis of proteins in both, academic protein structure studies as well as in process analytical applications. Here, the application of EC-QCL based IR transmission spectroscopy is introduced for i) monitoring of the protein secondary structure and ii) rapid screening of the thermal history of commercial milk samples without prior sample…preparation. Methods: Proteins present in milk were measured by QCL-IR and FTIR spectroscopy and spectra were compared. Dynamic conformational changes were followed by QCL-IR spectroscopy after chemical denaturation. Sixteen commercial milk samples were surveyed by QCL-IR spectroscopy and classified according to the experienced heat load during processing. Results: The 4–5 times higher applicable transmission path length (38 μ m for QCL-IR vs. 8 μ m for FTIR measurements) allows robust measurements of the protein amide I band in aqueous solutions. It was shown that IR spectra of the protein amide I band acquired by EC-QCL transmission spectroscopy are comparable to FTIR spectra and the acquired spectra were employed for the study of conformational changes in protein standard solutions. Furthermore, a classification analysis of commercial bovine milk samples based on their thermal history was accomplished. Conclusions: The potential application of EC-QCL IR spectroscopy was demonstrated as a tool for following conformational changes of the secondary protein structure as well as for fast screening to estimate the heat load applied to commercial milk.
Keywords: Quantum cascade laser, IR spectroscopy, protein secondary structure, milk analysis
Abstract: Background: Understanding the efficiency of a transdermal medical drug requires the characterization of its diffusion process, including its diffusion rate, pathways and physical state. Objective: The aim of this work is to develop a strategy to achieve this goal. Methods: FTIR spectroscopic imaging in conjunction with a Franz cell and HPLC measurements were used to examine the transdermal penetration of deuterated tert- butyl phenylchloroethylurea (tBCEU), a molecule with a potential anticancer action. tBCEU has been solubilized in an expedient solvent mixture and its diffusion in hairless mouse skin has been studied. Results: The results…indicate that tBCEU diffuses across the skin for more than 10 hours with a rate comparable to selegiline, an officially-approved transdermal drug. IR image analyses reveal that after 10 hours, tBCEU penetrates skin and that its spatial distribution does not correlate with neither the distribution of lipids nor proteins. tBCEU accumulates in cluster domains but overall low concentrations are found in skin. FTIR spectroscopic imaging additionally reveals that tBCEU is in a crystalline form. Conclusions: The results suggest that tBCEU is conveyed through the skin without preferential pathway. FTIR spectroscopic imaging and transdermal diffusion measurements appear as complementary techniques to investigate drug diffusion in skin.
Keywords: Infrared microspectroscopy, tert-butyl phenylchloroethylurea, diffusion pathway, diffusion kinetics, drug physical state, Franz cell, HPLC
Abstract: MD – the white areas on a mammogram (also known as breast density), has long been recognised as an indicator of breast cancer (BC) risk and mammographic masking. Recent legislation in 32 American states has mandated the inclusion of mammography density information in reports for women in the higher two MD quartiles, and it is a growing consideration world-wide. While the mammogram is currently the only means of estimating MD, it suffers from a number of limitations. These are related to the accumulation of low dose ionising radiation used in mammography that limits its repeated use, particularly in young women, women…with previous radiation exposure, those having undergone prior surgery, or those with radio-sensitising gene mutations. This review compares and contrasts the variety of emerging technologies that can provide a quantitative and true volumetric analysis of breast density, without the use of ionising radiation.
Keywords: Mammographic density, breast cancer risk factor, magnetic resonance imaging, fibroglandular tissue, ultrasound, ionising radiation
Abstract: Background: Open magnetic resonance (MRO) is an advanced technology where the configuration of the magnet allows images to be captured with patients sitting and standing as well as supine. This is relevant where posture and the effect of gravity have an impact on the region imaged; pelvic organ prolapse (POP) is an example. In POP, MRO provides standing images that allow detection of organ prolapse due to the effects of posture and gravity; and three-dimensional (3D) models generated from 2D images further enhance definition of the morphological features of POP. Purpose: To describe how high quality open MR…2D images can be modeled to create three dimensional images of the pelvic structures and enhance evaluation of patients with POP. Material and Methods: A cohort of 31 women (13 with POP and 18 asymptomatic controls) were recruited for MRO studies using a 0.5 T scanner. Axial and sagittal T2-weighted pelvic scans were obtained standing, sitting and supine. Manual segmentation using Analyze 12.0 software was then used to construct the 3D models. Results: The 3D modeling method developed generated images that were realistic and clearly revealed the extent of downward descent of pelvic structures in patients with POP. Generating 3 D images from sitting and standing 2D images enabled the extent and detail of organ descent to be better defined. Conclusion: A new protocol for open MR imaging combined with methodology for 3D image modeling of the female pelvis provides enhanced anatomic definition and will allow more comprehensive evaluation of the extent of POP due to the impact of posture and gravity.
Keywords: Open magnetic resonance imaging (MRO), pelvic organ prolapse (POP), three-dimensional (3D) modeling