PEGA supports for combinatorial peptide synthesis and solid-phase enzymatic library assays.
Permeable resins cross-linked with long PEG chains were synthesized for use in solid-phase enzyme library assays. High molecular weight bis-amino-polyethylene glycol (PEG) 4000, 6000, 8000 were synthesized by a three-step reaction starting from PEG-bis-OH. Macromonomers were synthesized by partial or di-acryloylation of bis-amino-PEG derivatives. Bis/mono-acrylamido-PEG were copolymerized along with acrylamide by inverse suspension copolymerization to yield a less cross-linked resin (Type I, compounds 6-9). Furthermore, acryloyl-sarcosin ethyl ester was co-polymerized along with bis-acrylamido PEG to obtain more crosslinked capacity resin (Type II, compounds 13-19). N,N-Dimethylacrylamide was used as a co-monomer in some cases. The polymer was usually obtained in a well-defined beaded form and was easy to handle under both wet and dry conditions. The supports showed good mechanical properties and were characterized by studying the swelling properties, size distribution of beads, and by estimating the amino group capacity. Depending on the PEG chain length, the monomer composition and the degree of cross-linking the PEGA supports showed a high degree of swelling in a broad range of solvents, including water, dichloromethane, DMF, acetonitril, THF and toluene: no swelling was observed in diethyl ether. The PEGA resins (Type I) with an amino acid group capacity between 0.07 and 1.0 mmol/g could be obtained by variation of the monomer composition in the polymerization mixture. Fluorescent quenched peptide libraries were synthesized on the new polymer using a multiple column library synthesizer and incubated with the matrix metalloproteinase MMP-9 after it had been activated by 4-aminophenyl mercuric acetate resulting in 67/83 kDa active enzyme. The bright beads were separated manually under a fluorescence microscope and sequenced to obtain peptide substrates for MMP-9. After treatment with ethylene diamine, high-loaded resins (Type II) have been employed in continuous flow peptide synthesis to yield peptides in excellent yield and purity.
Renil
M
M
Department of Chemistry, Carlsberg Laboratory, Valby, Denmark.
Ferreras
M
M
Delaisse
J M
JM
Foged
N T
NT
Meldal
M
M
D000081:Acetamides / Q000737:chemistry; D000178:Acrylamides / Q000737:chemistry*; D017364:Collagenases / Q000737:chemistry*; D009682:Magnetic Resonance Spectroscopy; D020780:Matrix Metalloproteinase 9; D019151:Peptide Library; D010455:Peptides / Q000138:chemical synthesis* / Q000737:chemistry; D011092:Polyethylene Glycols / Q000737:chemistry*; D014227:Triazines / Q000737:chemistry; D000099166:Polyacrylamides
D000081:Acetamides; D000178:Acrylamides; D017364:Collagenases; D020780:Matrix Metalloproteinase 9; D019151:Peptide Library; D010455:Peptides; D011092:Polyethylene Glycols; D014227:Triazines; C110274:2,4,6-mesitylenesulfonyl-3-nitro-1,2,4-triazolide; C113798:polyethyleneglycol-polyacrylamide copolymer; D000099166:Polyacrylamides; C014026:hexamethylene bisacetamide