The United States of America As Represented by the Department of Health

The United States of America

Constantine Londos

John J Egan

Alan R Kimmel

Andrew S Greenberg

The present invention provides isolated nucleic acid sequences, i.e., polynucleotides, which encode a family of perilipin proteins. The present invention also provides isolated, substantially purified perilipin proteins which are useful as markers for differentiating true adipocytes from non-adipocyte cells which, as a result of pathophysiological conditions, assume adipocyte characteristics and become lipid-laden. The present invention further provides methods for producing a substantially purified perilipin protein and methods for detecting the presence of such perilipin proteins in a biological samples.

Cloning of perilipin proteins

A method for assaying protein kinases that phosphorylate peptides such as Kemptide, such as cAMP-dependent protein kinase, or a glycogen synthase peptide, which is an excellent substrate for protein kinase C. Upon sequentially processing of reaction mixtures through tandem columns of cation and anion exchange resins improved separation of ATP from phosphorylated peptides is achieved such that radioactivity in background samples is nearly nil and the yield of phosphorylated peptides is high. This method is generally applicable to any protein kinase so long as the substrate peptide is appropriately structured such that the peptide retains a net positive charge when fully phosphorylated so that the peptide will adhere to the cation exchange resin and pass through the anion exchange resin. This method reduces labor, radioactivity, enzyme requirements, and costs of assaying protein kinases.

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Garrett Park, MD, United States of America

Constantine Londos