Positional scanning substrate combinatorial library (PS-SCL) approach to define caspase substrate specificity

Positional scanning substrate combinatorial library (PS-SCL) is a powerful tool for studying substrate specificity of proteolytic enzymes. Here, we describe the protocol for analyzing S4-S2 pockets preferences of caspases using PS-SCL. Additionally, we describe procedures for the identification of o...

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Bibliographic Details
Published inMethods in molecular biology (Clifton, N.J.) Vol. 1133; p. 41
Main Authors Poręba, Marcin, Szalek, Aleksandra, Kasperkiewicz, Paulina, Drąg, Marcin
Format Journal Article
LanguageEnglish
Published United States 2014
Subjects
Amino Acid Sequence
Caspases - chemistry
Caspases - genetics
Caspases - metabolism
Combinatorial Chemistry Techniques
Humans
Kinetics
Molecular Biology - methods
Peptide Library
Substrate Specificity
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Summary:Positional scanning substrate combinatorial library (PS-SCL) is a powerful tool for studying substrate specificity of proteolytic enzymes. Here, we describe the protocol for analyzing S4-S2 pockets preferences of caspases using PS-SCL. Additionally, we describe procedures for the identification of optimal substrates sequence after PS-SCL, solid phase synthesis, and purification of selected fluorogenic substrates, as well as their kinetic analysis.
ISSN:1940-6029
DOI:10.1007/978-1-4939-0357-3_2