Protocol abstract


Nature Protocols 2, 2126 - 2138 (2007)
Published online: 30 August 2007 | doi:10.1038/nprot.2007.305

Subject Categories: Genomics and proteomics | Isolation, purification and separation | Chemical modification | Synthetic chemistry | Pharmacology and toxicology | Biochemistry and protein analysis

Inhibitor fingerprinting of metalloproteases using microplate and microarray platforms: an enabling technology in Catalomics

Wei L Lee1, Junqi Li2, Mahesh Uttamchandani1, Hongyan Sun2 & Shao Q Yao1,2,3


One of the most fundamental properties of an enzyme is its selectivity, a property that has proved highly challenging to understand. Recent developments offer methodologies to rapidly establish activity-dependent profiles of enzymes. In this protocol, we describe methods to elucidate inhibitor fingerprints of enzymes. By taking advantage of well-defined small-molecule inhibitor libraries and the screening throughput offered from microplate and microarray platforms, we provide step-by-step application of the methodology toward the global characterization of metalloproteases, an important class of enzymes involved in numerous diseases and cellular processes. The same strategy is nonetheless applicable to virtually any given enzyme class, provided suitable experimental design and chemical inhibitor libraries are carefully implemented. We are able to routinely fingerprint as many as 2,000 independent enzyme interactions on the microplate platform within a span of approx7 h; however, the same throughput is attained within 5 h on the microarray platform.

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  1. Department of Biological Science, National University of Singapore, 3 Science Drive 3, Singapore 117543, Republic of Singapore.
  2. Department of Chemistry, National University of Singapore, 3 Science Drive 3, Singapore 117543, Republic of Singapore.
  3. NUS MedChem Program of the Office of Life Sciences, National University of Singapore, 3 Science Drive 3, Singapore 117543, Republic of Singapore.

Correspondence to: Shao Q Yao1,2,3 e-mail: chmyaosq@nus.edu.sg

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