Conference Publication Details
Mandatory Fields
Keyes, TE;Leane, D;Forster, RJ;Moran, N;Kenny, D
Opto-Ireland 2005: Optical Sensing and Spectroscopy
New insights into the molecular mechanisms of thrombosis from high resolution surface enhanced Raman microscopy
2005
January
Published
1
7 ()
Optional Fields
SILVER ELECTRODE SPECTROSCOPY CYSTEINE SPECTRA ALPHA(IIB)BETA(3) DISULFIDES
221
231
Hemostasis and Thrombosis refer respectively to the normal or pathological blood clotting within the vasculature. Occlusion of a blood vessel due to thrombosis can reduce or completely stop blood supply to different tissues or organs with the clinical consequences of myocardial infarction or stroke. Platelets are the cellular component which initiate thrombus formation, they activate in response to a variety of signals, such as interactions with a damaged blood vessel. alpha(IIb)beta(3) is a membrane bound integrin protein responsible for regulating adhesion of the activated platelet to damaged blood vessels. It exists in both activated and non-activated states displaying high and low affinity respectively for ligands such as fibrinogen. alpha(IIb)beta(3) determines the "stickiness" of the blood platelet and is therefore, a logical target for therapeutic measures to control thrombus formation. During the past decade considerable progress has been made to identify the role of the alpha(IIb)beta(3) complex in platelet-mediated thrombus formation and the structure of alpha(IIb)beta(3) has been extrapolated from the crystal structure of related integrins. However, despite these advances, the bimolecular mechanisms underlying the activation of alpha(IIb)beta(3) remain poorly understood. We have recently embarked on spectroscopic and photochemical studies of this protein in order to provide enhanced insight into the mechanism and dynamics of its activation. In this contribution, we describe methodologies of deriving surface enhanced Raman Spectroscopy of alpha(IIb)beta(3) on nanostructured metal surfaces, fabricated by a number of methods. We compare activation of alpha(IIb)beta(3) by SERS using a range of known activation conditions including Mn(II), EDTA and dithiotheritol (DTT). By studying the behaviour of the disulfide and CS marker vibrations in the spectral region 400 to 800 cm(-1) using SERS we confirm that activation results in significant conformational change in the protein, and most interestingly, that the response is not the same for every agonist. This mechanistic difference has implications for the biochemical study of this protein (and indeed for understanding the role of this integrin in response to different agonists).
10.1117/12.605333
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