Structure analysis and molecular recognition studies of bio-functionalized surfaces

Biological and chemical reactions involved in physiological processes are often complex and very sophisticated. Such processes can be mimicked in the laboratory to obtain important knowledge, valuable for the development of new diagnostic methods, drugs and biosensors. This thesis includes investigations of bio-functionalized surfaces that can be used as model systems to mimic naturally existing biomolecular recognition processes. In this thesis, three different peptides, of our own design, that mimic parts of the second and third intracellular loops of the α 2A-adrenergic receptor, are studied. The peptides were immobilized onto gold substrates, through thiol chemistry. The interaction between the peptides and the G-protein was investigated using surface plasmon resonance (SPR). The G-protein showed the highest binding capability for surfaces functionalized with a peptide mimicking the n-terminal of the third intracellular loop (GPR-i3n). The binding was enhanced when the pure GPR-i3n peptide was mixed with a short oligopeptide (3GC). A tentative explanation for the obtained results is that the presence of the 3GC molecule enables conformational changes of the GPR-i3n monolayer which affect the interaction with the G-protein. The results from the SPR measurements also indicated that the conformation of the G-protein was kept intact during the interaction with a peptide mimicking the c-terminal of the third intracellular loop (GPR-i3c). Multilayers were formed on the surfaces functionalized with a peptide mimicking the second intracellular loop (GPR-i2c) and the GPR-i3n peptide. We suggest that conformational changes of the G-protein are induced during the interaction with the surfaces functionalized with the GPR-i3n and GPR-i2c peptides. Comprehensive surface characterizations of four biomolecular systems, based upon the functional groups: noradrenaline, phenylboronic-ester, phenylboronic-acid and benzenesulfonamide, are presented in the thesis. The aim is to develop a platform for detailed molecular recognition studies on surfaces. The molecular systems were characterized using infrared spectroscopy, X-ray photoelectron spectroscopy, near edge X-ray absorption fine structure spectroscopy, ellipsometry and contact angle goniometry. Noradrenaline was chosen as it is a neurotransmitter that interacts with the extracellular loops of adrenergic receptors. In this work, the noradrenaline analogue (Nor-Pt) of our own design, was equipped with aSH handle to be linked to surfaces and with the free noradrenaline group available for interaction studies. The Nor-Pt molecules were organized on the surfaces with the sulfur atom close to the gold substrate and the aromatic ring available for possible interactions with other biomolecules in the ambient media. The main component of the C=O vibrational mode present …