Building a human kinase gene repository: Bioinformatics, molecular cloning, and functional validation

Jaehong Park, Yanhui Hu, T. V.S. Murthy, Fredrik Vannberg, Binghua Shen, Andreas Rolfs, Jessica E. Hutti, Lewis C. Cantley, Joshua LaBaer, Ed Harlow, Leonardo Brizuela

Research output: Contribution to journalArticle

36 Scopus citations

Abstract

Kinases catalyze the phosphorylation of proteins, lipids, sugars, nucleosides, and other important cellular metabolites and play key regulatory roles in all aspects of eukaryotic cell physiology. Here, we describe the mining of public databases to collect the sequence information of all identified human kinase genes and the cloning of the corresponding ORFs. We identified 663 genes, 511 encoding protein kinases, and 152 encoding nonprotein kinases. We describe the successful cloning and sequence verification of 270 of these genes. Subcloning of this gene set in mammalian expression vectors and their use in high-throughput cell-based screens allowed the validation of the clones at the level of expression and the identification of previously uncharacterized modulators of the survivin promoter. Moreover, expressions of the kinase genes in bacteria, followed by autophosphorylation assays, identified 21 protein kinases that showed autocatalytic activity. The work described here will facilitate the functional assaying of this important gene family in phenotypic screens and their use in biochemical and structural studies.

Original languageEnglish (US)
Pages (from-to)8114-8119
Number of pages6
JournalProceedings of the National Academy of Sciences of the United States of America
Volume102
Issue number23
DOIs
StatePublished - Jun 7 2005

Keywords

  • Autophosphorylation
  • Cell-based screens
  • High-throughput cloning
  • Kinome
  • Survivin

ASJC Scopus subject areas

  • General

Fingerprint Dive into the research topics of 'Building a human kinase gene repository: Bioinformatics, molecular cloning, and functional validation'. Together they form a unique fingerprint.

  • Cite this

    Park, J., Hu, Y., Murthy, T. V. S., Vannberg, F., Shen, B., Rolfs, A., Hutti, J. E., Cantley, L. C., LaBaer, J., Harlow, E., & Brizuela, L. (2005). Building a human kinase gene repository: Bioinformatics, molecular cloning, and functional validation. Proceedings of the National Academy of Sciences of the United States of America, 102(23), 8114-8119. https://doi.org/10.1073/pnas.0503141102