Research On Fmoc Chemistry Solid Phase Synthesis Of Difficult Peptide AM-55

Solid Phase Peptide Synthesis (SPPS) is an important way of obtaining peptides with good biological activity, which includes Fmoc Chemistry and Boc Chemistry. Fmoc Chemistry satisfies the requirement of rapid and auomatic peptide preparation, so it has greatly boosted the domains such as biochemistry, immunology, molecular microbiology and medicine, which all have a great demand in peptides.Fmoc Chemistry SPPS has become quite mature. But, for the synthesis of peptides which include many hydrophobic amino acids and formeβ-sheets easily, there are still difficulties. Repeated incomplete deprotection and coupling usually happen, generally deletion peptide and truncated peptide emerge with lower yield. These peptides are called difficult peptide. So, at the present time, it is urgent to solve the problem of the synthesis of difficult peptides.Commonly, there are two methods to solve the long peptide synthesis: Step-by-step SPPS (SSPPS) and Convergent SPPS(CSPPS). The former are usually used in synthesis of peptides containing less than 30 AA, though easily operation but low yields and poor purity. For longer peptide, the CSPPS is more suitable.We study a sequence AM-55 which may have the biological activity (ATALV ALVFS ILTGS ALGII GYGLL MAVTG ALEDE SLVEK ANKFW GIYST CDFIM). Fisrtly, we synthesized AM-55 via Step-by-step SPPS, and got a series of difficult peptides including H-IM-45-NH₂. Then we synthesized it by Convergent SPPS, and obtained four segments. At last, we explored how to couple them. The main products are listed as below:H-WM-11-NH₂ H-SM-20-OH H-GM-26-NH₂H-LM-31-OH H-GM-35-NH₂ H-AM-40-OHH-IM-45-NH₂ Fmoc-IM-9-OH Fmoc-AG-16-OHFmoc-SG-16-OH Fmoc-AG-14-OH H-IM-9-OH H-AG-16-OH H-SG-16-OH H-AG-14-OHWe had adopted Fmoc Chemistry SPPS to synthesize all the target molecules above, explored the synthetic method of difficult sequence peptides with different procedures, and compared the effects of various factors.Especially, we payed attention to:(1) The test stability of Sub;(2) The effects of Resins and Sub, Temperature and Coupling Reagents to Step-by-step SPPS of long difficult peptides;(3) We also explored the different conditions and optimized them to synthesize short difficult peptides;(4) How to couple the segments.Meanwhile, we had accomplished:(5) WM-11,SM-20,GM-26,LM-31,GM-35,AM-40,IM-45 were synthesized successfully via Step-by-step SPPS, in which the longest peptide is 45AA residues;(6) IM-9, AG-16, SG-16 and AG-14 had been synthesized successfully.Results indicated that:(7) The synthesis of AM-55 is difficult via Step-by-step SPPS;(8) Fmoc- IM-9 -OH and IM-9 can be synthesized easily with high purity because of their short length, and Fmoc-IM-9-OH can also be modified easily;(9) Fmoc-AG-16-OH,Fmoc-SG-16-OH and Fmoc-AG-14-OH can only be synthesized with low purity for their poor solubility; and their cleavage outcomes can't be purified and analyzed because of their dissolubility in water and methanol;(10) Fmoc-AG-16-OH, Fmoc-SG-16-OH and Fmoc-AG-14-OH can hardly be modified or coupled;(11) The key factor of synthesis of difficult peptides is solubility; the purity can be enhanced by higher temperature and additive;(12) The synthesis of AM-55 is also difficult even though Convergent SPPS is used.This study can be used for reference when you want to synthesize other difficult peptides.