Investigations Of Proton Pumping Behaviors Of Retinal Proteins And Corresponding Material Techniques In Potential Photoactive Devices

As a retinal protein,bacteriorhodopsin(BR)was discovered in the membrane of halobacterium salinarum living in the salty lakes.Another retinal protein, archaerhodopsin 4(AR4)was found by researchers from Fudan University from Tibet.These proteins contain a retinal molecule as chrome and exhibit a unique photocycle leading to a proton pumping function.The latest three decades have witnessed much progress of BR researches,and even the conceptual assumption of 'protein computer' was put forward.The retinal protein has been one of the most attractive proteins,especially the most well-known membrane protein.Nevertheless,further studies on the retinal proteins are still required,basically along the two lines.One is the fundamental study on their structure-function relationship.The protein is the result of millions of years' evolution.It is hard to fully understand how such a simple structure can have so complicated functions.The other is the technical applications of retinal proteins in the photochromic and photoelectric fields.The special functions of photocycle and proton pump and the especially stable structure amongst active proteins encourages the associated investigations towards potential applications.To date,some models in data storage and anti-counterfeiting etc.have been suggested.This Ph.D thesis concerns an interdisciplinary research among fields of polymer materials,life science,and information science.The main content of this thesis is to study the kinetics of proton transfer in the photocyle,to examine the self assembly behaviors of retinal proteins,to fabricate microarrays with oriented proteins as a photoreceptor,and to prepare polymer-protein composites as potential information materials.The mail creative investigations are summarized as follows:(1)A photoelectrochemical approach was put forward to detect pK_a of protein release complex(PRC)of retinal rpoteins at the photo-excited state.By detecting photoinduced potentials that originate from the proton concentration changes(â–³H+) generated by proteins near the indium tin oxide(ITO) electrode,we show that the kinetics of release and uptake can be followed in a broad pH range,and the pK_a of PRC can be easily determined under different conditions.A kinetic analysis was performed and a formula describing proton fluxes of wild-type BR and AR4,and D96N mutant of BR was derived.The pH dependence of the fraction of fast release over the whole release was used to determine the pK_a for proton release in the photocycles of these retinal proteins.Measurements can also be performed in conventional buffer solutions and crown ether,which is not available before the invention of this approach.This feature enables us to study effects of chemicals with high buffer capacity,and significant effects of buffers and crown ether on proton pumping behaviors of retinal proteins were revealed.This method is a powerful tool for investigation of light-driven proton pumping mechanisms in a wide pH range.(2) Material techniques for photoelectric and photochromic BR devices were developed.(A) A microarray of orientated BR proteins with separated pixels was fabricated.A microarray protein chip was also prepared based on the technique of biology,chemistry and physics to be applied as a photoreceptor.(B) Further more, we invent a new method to prepare functional polymer/protein materials.Different from the previous way,a crosslinked super molecule network including active protein and polymer was prepared.The new material can provide a good distribution of protein molecules and a high stability.(3) Self assembly behaviors of retinal proteins were examined.Proteins were assembled into liposomes with prefeered orientation,and the self assembly behaviors of BR and AR4 were investigated in a comparative way.PEGylation of BR was also performed,and its influence on photoresponse and orientation of BR were examined.