Spectroscopic Study Of Conformational Changes Of Proteins And Polypeptide Under Multi-factors

Protein is the foundation of living matters and plays a very vital role in various life activities.The function of protein is directly related to its structure.The structure of proteins can be distinguished for four levels,among which the secondary structure is of great significance.The physiological functions of proteins and the interaction of proteins with other molecules are realized under certain solvent conditions,so it is valuable to study the structure changes in protein solution.The main methods for determining the secondary structure of proteins in solution are Circular Dichroism(CD),Nuclear Magnetic Resonance(NMR)and Fourier Transform Infrared Spectroscopy(FTIR),the development and application of the three methods are reviewed in this paper.However,these methods have high requirements for samples,and can not get information in situ and continuously.Attenuated Total Reflection Fourier Transform Infrared Spectroscopy(ATR-FTIR)is a non-destructive surface analysis technique that greatly reduces the effect of water on the spectra,making the spectra more realistic and reliable.In this paper,three different materials(trypsin,bovine serum albumin,polypeptide)was used to study the changes of Amide bonds in different states,different metal ions and different surfaces by ATR-FTIR technique,and then analyzed the element of protein secondary structure under different conditions.After trypsin,BSA and polypeptide powders dissolved in water,hydrogen bonds formed with water molecule,which promoted the rearranged of electron cloud density on the C=O and NH groups,resulting in Amide I and Amide II bands shifted.Changes of Amide bonds indicated that the three samples have different secondary structural components in two states.We proposed that the formation of proteins aqueous solution or the contact between the protein solution and surface were the main cause of this change,so we continued to explore the effects of different concentrations of metal ions(calcium cations,zinc cations)on the protein solution.The changed of trypsin on different surfaces(Zn Se and titanium dioxide)were also studied.The addition of different concentrations of calcium or zinc cations in the trypsin solution further caused the shift of the Amide I and Amide II bands,indicating that trypsin interacts with the hydrated metal cations and changes the secondary structure of the protein.On the surface of titanium dioxide,the shift of Amide I and Amide II bands indicated that the adhesion process of trypsin on different surfaces have different secondary structural element,and the quantity of trypsin on titanium dioxide surface was 20% more than on the Zn Se surface.Using curve fitting mathematical method to analysis the differential spectra,and quantitatively obtain the protein secondary structure component.The changes of shape and shift of Amide I and Amide II bands were further induced by adding different concentrations of calcium cations in BSA aqueous solution,and the larger concentration of calcium cations,the greater the changes of Amide bands.Curve fitting method was used to analyze the FTIR differential spectra of obtained by two different methods of subtracting background.The change trend of the secondary structure was the same,and the conclusion was consistent with the results obtained by Raman spectroscopy.Different concentrations of calcium or zinc cations were added to polypeptide aqueous solution,the Amide I band shifted to low wavenumber,and the Amide II band shifted to high wavenumber.It was confirmed that the addition of hydrated metal cations would lead to the changes of Amide bands,resulting in the secondary structure changes of polypeptide,and the change trend of the two metal cations was the same.Based on the experimental results,the thesis initially revealed the factors effecting the secondary structure of proteins in aqueous solution.Hydrogen bonds and surface interactions played significant roles in secondary structure changes of proteins.