Limiting racemization and aspartimide formation in microwave-enhanced Fmoc solid phase peptide synthesis.
Microwave energy represents an efficient manner to accelerate both the deprotection and coupling reactions in 9-fluorenylmethyloxycarbonyl (Fmoc) solid phase peptide synthesis (SPPS). Typical SPPS side reactions including racemization and aspartimide formation can occur with microwave energy but can easily be controlled by routine use of optimized methods. Cysteine, histidine, and aspartic acid were susceptible to racemization during microwave SPPS of a model 20mer peptide containing all 20 natural amino acids. Lowering the microwave coupling temperature from 80 degrees C to 50 degrees C limited racemization of histidine and cysteine. Additionally, coupling of both histidine and cysteine can be performed conventionally while the rest of the peptide is synthesized using microwave without any deleterious effect, as racemization during the coupling reaction was limited to the activated ester state of the amino acids up to 80 degrees C. Use of the hindered amine, collidine, in the coupling reaction also minimized formation of D-cysteine. Aspartimide formation and subsequent racemization of aspartic acid was reduced by the addition of HOBt to the deprotection solution and/or use of piperazine in place of piperidine.
Palasek
Stacey A
SA
Bioscience Division, CEM Corporation, Matthews, North Carolina 28106, USA.
Cox
Zachary J
ZJ
Collins
Jonathan M
JM
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000737:chemistry*; D001224:Aspartic Acid / Q000031:analogs & derivatives* / Q000138:chemical synthesis; D005449:Fluorenes / Q000737:chemistry*; D007536:Isomerism; D008872:Microwaves; D010455:Peptides / Q000138:chemical synthesis*
D000596:Amino Acids; D005449:Fluorenes; C016456:N(alpha)-fluorenylmethyloxycarbonylamino acids; D010455:Peptides; D001224:Aspartic Acid; C075871:aspartimide