Effect Of Cholesterol On Human Islet Amyloid Polypeptide Interaction With Phospholipid Membranes

Amyloid protein is prone to misfolding and assembling to form ?-sheet-rich fibrous deposits under pathological conditions.Various diseases such as Alzheimer's disease(AD),type II diabetes(T2DM),Bovine Spongiform Encephalopathy(BSE),Parkinson's disease(PD)are related to the deposits of amyloid aggregations,consequently,the associated amyloid proteins have been concerned and researched widely by the scientists.Type II diabetes(T2DM)poses a significant threat to human health,and the deposition of human islet amyloid polypeptide(h IAPP)in diabetic patients may be an important cause of diabetes.h IAPP is composed of 37 amino acids co-secreted with insulin in islet ?-cells.The formation of oligomeric intermediates during the process of h IAPP aggregation is considered to be a major component resulting in the destruction of the cell membrane.It is found that the interaction of h IAPP with the membrane is a critical step in the destruction of the membrane,whereas the N-terminal peptide hIAPP1-19 is the major region of the peptide-membrane interaction.Human IAPP1-19 has little fibrosis tendency in water or in membrane,maintaining a relatively stable structure,but can cause membrane damage similarly to the full length hIAPP in part.For this reason,it is an excellent model peptide to study hIAPP interaction with membrane.Cholesterol is an important component of the cell membrane,and forms lipid rafts with sphingomyelin on the membrane which can change the membrane properties such as fluidity,thickness and rigidity markedly.High cholesterol levels of diabetic patients may have a certain impact on the interaction between h IAPP and islet ?-cell membrane.Furthermore,the amino acid sequence RLANFLV from position 11 to 17 of hIAPP1-19 satisfies the inverted cholesterol recognition/interaction amino-acid consensus sequence(CARC),thus,it may be conducive to hIAPP tendentious binding to cholesterol.In the CARC-like sequence,phenylalanine(F)may play an important role for recognition of peptide and cholesterol.Based on the importance of h IAPP1-19 in the process of hIAPP interacting with membrane,this paper mainly studied the interactions of h IAPP1-19 and its F15 L mutant hIAPP1-19/F15 L with different phospholipid model membranes and explored the effects of cholesterol on the peptide-membrane interactions using circular dichroism(CD),~1H-NMR,31P-NMR,ion leakage and SDS-PAGE experiments.In POPC/POPG(8:2)vesicle system,the interaction between hIAPP1-19 and membrane is stronger with peptide inserting into lipids.There are almost no difference beween structures,membrane binding strengths or destruction abilities of the two peptides in the absence of cholesterol.The adition of cholesterol in mixed membranes increases the amount of ?-helical structure,enhances the binding of h IAPP1-19 to the membrane,and deteriorates the damage of the membrane.In contrast,these properties and/or behaviors of hIAPP1-19/F15 L are less affected by cholesterol.Our results suggest that hIAPP1-19 may have a direct binding to cholesterol to some extent and F15 may be involved in the interaction of hIAPP1-19 with cholesterol.The substitution of F15 by L could weaken the recognition of the peptide to cholesterol,leading to a decrease in peptide-membrane interaction and membrane damage.In DOPC/DPPC(1:2)vesicle system,h IAPP1-19 and h IAPP1-19/F15 L are mainly present as a random coil structure.The binding constant of peptide to the zwitterionic membrane is reduced by about two orders of magnitude compared with the membrane containing anionic component(POPC/POPG 8:2),and the destruction of the phospholipid membrane induced by the peptides is relatively small.When DOPC,DPPC and Chol are mixed according to the ratio of 1:2:1,cholesterol could form a lipid raft on the membrane.Although the presence of cholesterol does not have a significant effect on the structure of the two peptides,it promotes the destruction of the membrane induced by h IAPP1-19,but has little effect on the membrane destruction induced by hIAPP1-19/F15 L.It can be hypothesized that the structure of peptide may not have an absolute correlation with its ability destroying membrane.The study on the interaction of hIAPP1-19 with membrane and the role of cholesterol in the process are important for understanding the binding mode of hIAPP to phospholipid membrane and the cytotoxicity mechanism of the amyloid peptide.