PNA-encoded protease substrate microarrays.
Our current understanding of the role and regulation of protease activity in normal and pathogenic processes is limited by our ability to measure and deconvolute their enzymatic activity. To address this limitation, an approach was developed that utilizes rhodamine-based fluorogenic substrates encoded with PNA tags. The PNA tags address each of the substrates to a predefined location on an oligonucleotide microarray through hybridization, thus allowing the deconvolution of multiple signals from a solution. A library of 192 protease substrates was prepared by split and mix combinatorial synthesis. The methodology and validation of this approach for profiling proteolytic activity from single proteases and from those in crude cell lysates as well as clinical blood samples is described.
Institut de Science et d'Ingénierie Supramoléculaires, Université Louis Pasteur, 8 allée Guaspard Monge, 67000 Strasbourg, France. winssinger@isis-ulp.org
Nicolas
N
Winssinger
Robert
R
Damoiseaux
David C
DC
Tully
Bernhard H
BH
Geierstanger
Keith
K
Burdick
Jennifer L
JL
Harris
D006801:Humans; D006868:Hydrolysis; D019169:Jurkat Cells; D020411:Oligonucleotide Array Sequence Analysis / Q000379:methods*; D010447:Peptide Hydrolases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D019151:Peptide Library; D020135:Peptide Nucleic Acids / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D040901:Proteomics / Q000379:methods; D013379:Substrate Specificity / Q000235:genetics
D019151:Peptide Library; D020135:Peptide Nucleic Acids; D010447:Peptide Hydrolases