Dataset Information

Engineering Pak1 Allosteric Switches.

ABSTRACT: P21-activated kinases (PAKs) are important regulators of cell motility and morphology. It has been challenging to interrogate their functions because cells adapt to genetic manipulation of PAK, and because inhibitors act on multiple PAK isoforms. Here we describe genetically encoded PAK1 analogues that can be selectively activated by the membrane-permeable small molecule rapamycin. An engineered domain inserted away from the active site responds to rapamycin to allosterically control activity of the PAK1 isoform. To examine the mechanism of rapamycin-induced PAK1 activation, we used molecular dynamics with graph theory to predict amino acids involved in allosteric communication with the active site. This analysis revealed allosteric pathways that were exploited to generate kinase switches. Activation of PAK1 resulted in transient cell spreading in metastatic breast cancer cells, and long-term dendritic spine enlargement in mouse hippocampal CA1 neurons.

SUBMITTER: Dagliyan O

PROVIDER: S-EPMC5562282 | biostudies-literature | 2017 Jul

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

Engineering Pak1 Allosteric Switches.

Dagliyan Onur O Karginov Andrei V AV Yagishita Sho S Gale Madeline E ME Wang Hui H DerMardirossian Celine C Wells Claire M CM Dokholyan Nikolay V NV Kasai Haruo H Hahn Klaus M KM

ACS synthetic biology 20170406 7

P21-activated kinases (PAKs) are important regulators of cell motility and morphology. It has been challenging to interrogate their functions because cells adapt to genetic manipulation of PAK, and because inhibitors act on multiple PAK isoforms. Here we describe genetically encoded PAK1 analogues that can be selectively activated by the membrane-permeable small molecule rapamycin. An engineered domain inserted away from the active site responds to rapamycin to allosterically control activity of ...[more]

PMID: 28365983

Similar Datasets

Project description:Proteins are dynamic molecules that undergo conformational changes to a broad spectrum of different excited states. Unfortunately, the small populations of these states make it difficult to determine their structures or functional implications. Computer simulations are an increasingly powerful means to identify and characterize functionally relevant excited states. However, this advance has uncovered a further challenge: it can be extremely difficult to identify the most salient features of large simulation data sets. We reasoned that many functionally relevant conformational changes are likely to involve large, cooperative changes to the surfaces that are available to interact with potential binding partners. To examine this hypothesis, we introduce a method that returns a prioritized list of potentially functional conformational changes by segmenting protein structures into clusters of residues that undergo cooperative changes in their solvent exposure, along with the hierarchy of interactions between these groups. We term these groups exposons to distinguish them from other types of clusters that arise in this analysis and others. We demonstrate, using three different model systems, that this method identifies experimentally validated and functionally relevant conformational changes, including conformational switches, allosteric coupling, and cryptic pockets. Our results suggest that key functional sites are hubs in the network of exposons. As a further test of the predictive power of this approach, we apply it to discover cryptic allosteric sites in two different ?-lactamase enzymes that are widespread sources of antibiotic resistance. Experimental tests confirm our predictions for both systems. Importantly, we provide the first evidence, to our knowledge, for a cryptic allosteric site in CTX-M-9 ?-lactamase. Experimentally testing this prediction did not require any mutations and revealed that this site exerts the most potent allosteric control over activity of any pockets found in ?-lactamases to date. Discovery of a similar pocket that was previously overlooked in the well-studied TEM-1 ?-lactamase demonstrates the utility of exposons.

Dataset Information

Engineering Pak1 Allosteric Switches.

Publications

Engineering Pak1 Allosteric Switches.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets