Positional scanning peptide libraries for kinase substrate specificity determinations: straightforward and reproducible synthesis using pentafluorophenyl esters.
An efficient method to synthesize positional scanning synthetic combinatorial libraries (PS-SCLs) for studying the specificity of protein kinases is presented. Isokinetic ratios for pentafluorophenyl esters were determined iteratively using a new approach incorporating high performance liquid chromatography (HPLC) quantification and statistical experimental design. In the development process a large amount of work was put in to find efficient ways of screening for new isokinetic mixtures and to optimize the process of PS-SCL synthesis. The newly developed methods for the screening of isokinetic mixtures could be used for the screening of other interesting mixtures, but more importantly, the isokinetic ratios determined for the preactivated pentafluorophenyl esters were incorporated into a new efficient protocol. This straightforward protocol allows for a convenient synthesis of high quality PS-SCLs regardless of previous experience in solid phase synthesis.
Ljungdahl
Thomas
T
Department of Chemistry, Medicinal Chemistry, University of Gothenburg, Göteborg, Sweden.
Veide-Vilg
Jenny
J
Wallner
Fredrik
F
Tamás
Markus J
MJ
Grøtli
Morten
M
D002851:Chromatography, High Pressure Liquid; D020650:Combinatorial Chemistry Techniques; D004952:Esters / Q000138:chemical synthesis* / Q000737:chemistry; D005464:Fluorobenzenes / Q000138:chemical synthesis* / Q000737:chemistry; D057166:High-Throughput Screening Assays; D015394:Molecular Structure; D019151:Peptide Library; D011494:Protein Kinases / Q000737:chemistry* / Q000378:metabolism; D013237:Stereoisomerism; D013379:Substrate Specificity
D004952:Esters; D005464:Fluorobenzenes; D019151:Peptide Library; D011494:Protein Kinases