Improved efficiency sos recruitment system: expression of the mammalian GAP reduces isolation of Ras GTPase false positives.
The Sos recruitment system (SRS) is a novel genetic method for detecting protein-protein interactions. The method is based on localizing Sos, a Ras guanyl nucleotide exchange factor (GEF), to the plasma membrane through interaction between two fusion proteins. Mammalian Ras can bypass the requirement for a functional Ras GEF and represents a predictable false positive in this system. This report demonstrates that introduction of mammalian GTPase activating protein (mGAP) reduces the isolation of Ras false positives in SRS screens of mammalian cDNA libraries, thereby significantly enhancing the efficiency of the system.
Aronheim
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Department of Molecular Genetics, The B. Rappaport Faculty of Medicine, PO Box 9649, Bat Galim, Haifa 31096, Israel.
D000818:Animals; D018797:Cell Cycle Proteins / Q000378:metabolism*; D003001:Cloning, Molecular / Q000379:methods*; D018076:DNA, Complementary / Q000235:genetics; D020690:GTPase-Activating Proteins; D015723:Gene Library; D008322:Mammals; D008565:Membrane Proteins / Q000378:metabolism*; D010749:Phosphoprotein Phosphatases / Q000378:metabolism*; D011485:Protein Binding; D011506:Proteins / Q000378:metabolism*; D012441:Saccharomyces cerevisiae; D020664:Son of Sevenless Proteins; D013379:Substrate Specificity; D020703:ras GTPase-Activating Proteins; D018631:ras Proteins / Q000378:metabolism*; D020666:ras-GRF1
D018797:Cell Cycle Proteins; D018076:DNA, Complementary; D020690:GTPase-Activating Proteins; D008565:Membrane Proteins; D011506:Proteins; D020664:Son of Sevenless Proteins; D020703:ras GTPase-Activating Proteins; D020666:ras-GRF1; D010749:Phosphoprotein Phosphatases; D018631:ras Proteins