Measurement of nitric-oxide-modified proteins by cavity ring-down spectroscopy

一氧化氮合酶产物通过与金属蛋白中的血红素中心相互作用发出信号，如鸟苷酸环化酶，并通过与蛋白质半胱氨酸硫醇的共价反应。这些亚硝基化反应在健康和疾病的真核细胞生物学中具有广泛的相关性。然而，亚硝基化蛋白经常被证明难以用临床和研究样品所需的灵敏度和特异性检测。我们正在开发空腔衰荡光谱来测量这些蛋白质修饰。我们相信，这项技术将大大提高灵敏度和特异性，它有可能彻底改变这一领域。在这里，我们提出设计和制造一种改进的具有样品光解界面的腔衰荡仪器。该仪器将采用先进的光谱学技术来测量生物标本中的S-亚硝基硫醇。它将被用于研究广泛的疾病和器官系统。它将比以前的技术更强大，更精确，在pM浓度下精确测量no修饰的蛋白质，同时最小限度地破坏样品。在第二个目标，它也将应用于蛋白质组微阵列快速，高通量的发现。

DESCRIPTION (provided by applicant) Nitric oxide synthase products signal through interactions with heme centers in metalloproteins, such as guanylate cyclase and through covalent reactions with protein cysteine thiols. These nitrosylation reactions have broad relevance to eukaryotic cell biology in health and disease. However, nitrosylated proteins have often proven difficult to detect with the sensitivity and specificity needed for clinical and reseach samples. We are developing cavity ring-down spectroscopy to measure these protein modifications. We believe that this technology will dramatically improve both sensitivity and specificity it has the potential to will revolutionize the field. Here, we propose to design and fabricate an improved cavity ring-down instrument with a sample photolysis interface. This instrument will use advanced optical spectroscopy techniques to measure S- nitrosothiols in biological specimens. It will be used to studying a broad range of diseases and organ systems. It will be more powerful and more precise than previous technologies, measuring NO-modified proteins accurately in pM concentrations with minimal sample disruption. In the second Aim, it will also be applied to proteomic microarrays for rapid, high-throughput discovery.