An entirely specific type I A-kinase anchoring protein that can sequester two molecules of protein kinase A at mitochondria

Christopher K. Means, Birgitte Lygren, Lorene K. Langeberg, Ankur Jain, Rose E. Dixon, Amanda L. Vega, Matthew G. Gold, Susanna Petrosyan, Susan S. Taylor, Anne N. Murphy, Taekjip Ha, Luis F. Santana, Kjetil Tasken, John D. Scott

Research output: Contribution to journalArticlepeer-review


A-kinase anchoring proteins (AKAPs) tether the cAMP-dependent protein kinase (PKA) to intracellular sites where they preferentially phosphorylate target substrates. Most AKAPs exhibit nanomolar affinity for the regulatory (RII) subunit of the type II PKA holoenzyme, whereas dual-specificity anchoring proteins also bind the type I (RI) regulatory subunit of PKA with 10-100-fold lower affinity. A range of cellular, biochemical, biophysical, and genetic approaches comprehensively establish that sphingosine kinase interacting protein (SKIP) is a truly type I-specific AKAP. Mapping studies located anchoring sites between residues 925-949 and 1,140-1,175 of SKIP that bind RI with dissociation constants of 73 and 774 nM, respectively. Molecular modeling and site-directed mutagenesis approaches identify Phe 929 and Tyr 1,151 as RI-selective binding determinants in each anchoring site. SKIP complexes exist in different states of RI-occupancy as single-molecule pulldown photobleaching experiments show that 41 ± 10% of SKIP sequesters two YFP-RI dimers, whereas 59 ± 10% of the anchoring protein binds a single YFP-RI dimer. Imaging, proteomic analysis, and subcellular fractionation experiments reveal that SKIP is enriched at the inner mitochondrial membrane where it associates with a prominent PKA substrate, the coiled-coil helix protein ChChd3.

Original languageEnglish (US)
Pages (from-to)E1227-E1235
JournalProceedings of the National Academy of Sciences of the United States of America
Issue number48
StatePublished - Nov 29 2011

ASJC Scopus subject areas

  • General


Dive into the research topics of 'An entirely specific type I A-kinase anchoring protein that can sequester two molecules of protein kinase A at mitochondria'. Together they form a unique fingerprint.

Cite this