Bioorganic Studies of Retinal Binding Proteins

描述（由申请人提供）：蛋白质-底物相互作用是支配信号传导途径和控制生理反应的生化事件的核心。这些相互作用决定了酶促反应的特异性，并且是控制生物反应的核心，尽管是由内源性化合物或药物诱导的。对蛋白质-底物相互作用的完整和透彻的理解是生物过程研究的基础。作为一个模型，我们已经进行了全面的研究，在视紫红质的波长调节。这是蛋白质-底物相互作用的一个显著例子，其中不同的蛋白质从相同的底物（即11-顺式-视黄醛）引发不同的性质。我们研究的目的是了解不同氨基酸残基与视网膜质子化席夫碱相互作用的影响，最终导致不同色素的不同波长最大值。我们提出了一种新的策略来研究这种独特的现象，基于在蛋白质环境中产生11-顺式-视网膜的质子化席夫碱，我们将工程师模仿视紫红质，然而，具有在手的优势，他们的三维图片。这将使我们能够合理地提出突变的氨基酸残基密切接触的结合基板和探测波长调节的起源。我们还从事13 C标记的视网膜的合成，以探测波长调节的构象方面。我们的第二个目标是开发这些视紫红质蛋白模拟物作为比色蛋白融合标签，这是生物技术领域高度期望的。我们计划证明我们的视紫红质蛋白模拟物作为理想的蛋白融合标签的实用性，其具有小尺寸、大结合腔、可调谐波长和开/关激活;即，加入发色团会使颜色变亮。所提出的蛋白质的小而可溶的性质以及它们独特的13桶腔沿着应允许结合各种发色团。通过各种结合的发色团和/或调节蛋白质与结合的发色团的相互作用来改变色素光谱特性的能力可以产生各种着色的蛋白质标签。

DESCRIPTION (provided by applicant): Protein-substrate interactions are at the heart of biochemical events that govern signaling pathways and control physiological responses. These interactions dictate specificity in enzymatic reactions and are central to the control of biological responses, albeit, induced by endogenous compounds or drugs. A complete and thorough understanding of protein-substrate interactions is fundamental to the study of biological processes. As a model, we have undertaken a comprehensive study of wavelength regulation in rhodopsin. This is a remarkable example of protein-substrate interaction in which different proteins elicit different properties from the same substrate, namely 11-cis-retinal. The aim of our research is to understand the effect of different amino acid residues interacting with the retinal protonated Schiff base that ultimately lead to differing wavelength maxima of different pigments. We propose a novel strategy to study this unique phenomenon based on generating protonated Schiff base of 11-cis-retinal in protein environments that we will engineer to mimic rhodopsin, however, with the advantage of having at hand their 3-dimensional picture. This will allow us to rationally propose mutagenesis of amino acid residues in close contact with the bound substrate and probe the origins of wavelength regulation. We are also engaged in the synthesis of 13C-labeled retinals to probe the conformational aspects of wavelength regulation. Our second aim proposes to develop these rhodopsin protein mimics as colorimetric protein fusion tags, which are highly desired by the biotechnology field. We plan to demonstrate the utility of our rhodopsin protein mimics as ideal protein fusion tags with their small size, large binding cavity, tunable wavelength, and on/off activation; i.e., addition of chromophore will turn on the color. The small and soluble nature of the proposed proteins along with their unique 13-barrel cavity should allow binding of a variety of chromophores. The ability to vary the pigments spectral characteristics, either through various bound chromophores and/or tuning of the protein interactions with the bound chromophore, can lead to a variety of pigmented protein tags.