Construction Of An Artificial Signal Transduction System On Bilayer Vesicles

Two artificial signal transduction systems,inspired by natural biological cell signal transduction,were constructed through special molecular recognition by NADH-dependent lactate dehydrogenase(LDH)as an effector and synthetic molecules as an artificial receptor incorparated in artificial bilayer membranes under certain conditions.In the present work,two cationic peptide lipids were synthesized, N,N-dihexadecyl-N^α-[6-(trimethylammonio)hexanoyl]alaninamide bromide(N⁺C₅Ala2C₁₆) prepared from 1-hexadecylamine and 1-bromohexadecane through five steps including substitution and condensation reactions,and 1-N-Hexadecyl-2-methyl-3-hydroxy-4-formyl-5-hydroxymethylpyridoxal chloride(PLC₁₆⁺),a derivertive of vitamine B₆,prepared from pyridoxal hydrochloride and 1-iodohexadecane as starting materials through four steps including deacetalization and condensation reactions.The total yields were 4.75%and 8.13%, respectively.The first artificial signal transduction system was constructed,employing PLC₁₆⁺ as receptor,GABA as an external signal and Cu(â…¡)ion as a mediator on bilayer membranes formed with N⁺C₅Ala2C₁₆at pH 8.00 and 40℃which was determined through investigating the complex formation of each of the four amino compounds Gly,β-Ala,GABA,Asp with PLC₁₆⁺,and the binding of the resulting complex with Cu²⁺in buffer solution at different pH. In the system,GABA formed a signal-receptor complex with the receptor through Schiff-base reaction at GABA concentration of 40mM on the surface of the mixing vesicles formed at a molar ratio of 20:1(N⁺C₅Ala2C₁₆:PLC_16<sup>+;and the resulting complex toward Cu(â…¡)ions in 2:1 stoichiometry was formed.As a result,the binding of the Cu(â…¡)ions with the complex led to 45%recovery of LDH relative activity on the surface of the vesicle.The second artificial signal transduetion system was constructed,employing N²,N^2'-2,5,8,11-Tetraoxadodecanedioyl-bis(N,N-dihexadecyl-L-histidinamide) [TeoBis(His2C₁₆)]as a receptor containing a histidine residue and a moiety of pseudo-crown ether,prepared through three steps,including substitution and condensation reaction at a yield of 51%,which was incorparated in bilayer membrane formed with N⁺C₅Ala2C₁₆at pH 7.00 and 40℃.The relative activity of LDH returned to 40.9%through preventing Cu²⁺ion inhibiting the enzymatic activity due to the formation of the complex between imidazole of TeoBis(His₂C₁₆)and Cu(â…¡)ion as an inhibitor.In the presence of K⁺,Na⁺,Li⁺ Rb⁺ or Cs⁺, the enzymatic activity was further enhanced due to the more effective binding of the receptor with Cu(â…¡)ion by a spatial effect through ineorparation of the alkali matel cation into the cavity site of pseudo-crown ether moity of the receptor TeoBis(His₂C₁₆).Among these alkali matel ion,K⁺ recovered the highest degree 85.9%of the relative activity of LDH.The novel artificial signal transduetion system employing a gemini lipid with histidine residues as receptor.