TABLE 1:
Information on protein expression constructs used in this study.
| Proteins | Sequence information | Notes |
|---|---|---|
| Nephrin(WT) | MALGTTLRASLLLLGLLTEGLAQLAIPASMEQKLISEEDLNNGSGGSGTGSGGSPGLHQPSGEPEDQLPTEPPSGPSGLPLLPVLFALGGLLLLSNASCVGGVLWQRRLRRLAEGISEKTEAGSEEDRVRNEYEESQWTGERDTQSSTVSTTEAEPYYRSLRDFSPQLPPTQEEVSYSRGFTGEDEDMAFPGHLYDEVERTYPPSGAWGPLYDEVQMGPWDLHWPEDTYQDPRGIYDQVAGDLDTLEPDSLPFELRGHLV | Nephrin plasma membrane targeting signal peptide (residues 1–29)—myc tag–Linker (LNNGSGGSGTGSGGS)—nephrin intracellular domain after Fibronectin type-III domain (residues 1036–1241) |
| Nephrin(Y3F) | MALGTTLRASLLLLGLLTEGLAQLAIPASMEQKLISEEDLNNGSGGSGTGSGGSPGLHQPSGEPEDQLPTEPPSGPSGLPLLPVLFALGGLLLLSNASCVGGVLWQRRLRRLAEGISEKTEAGSEEDRVRNEYEESQWTGERDTQSSTVSTTEAEPYYRSLRDFSPQLPPTQEEVSYSRGFTGEDEDMAFPGHLFDEVERTYPPSGAWGPLFDEVQMGPWDLHWPEDTYQDPRGIFDQVAGDLDTLEPDSLPFELRGHLV | Mutations: Y1176F, Y1193F, Y1217F |
| Nephrin(Y5F) | MALGTTLRASLLLLGLLTEGLAQLAIPASMEQKLISEEDLNNGSGGSGTGSGGSPGLHQPSGEPEDQLPTEPPSGPSGLPLLPVLFALGGLLLLSNASCVGGVLWQRRLRRLAEGISEKTEAGSEEDRVRNEYEESQWTGERDTQSSTVSTTEAEPYYRSLRDFSPQLPPTQEEVSYSRGFTGEDEDMAFPGHLFDEVERTFPPSGAWGPLFDEVQMGPWDLHWPEDTFQDPRGIFDQVAGDLDTLEPDSLPFELRGHLV | Mutations: Y1176F, Y1183F, Y1193F, Y1210F, Y1217F |
| Nck1 | MAEEVVVVAKFDYVAQQEQELDIKKNERLWLLDDSKSWWRVRNSMNKTGFVPSNYVERKNSARKASIVKNLKDTLGIGKVKRKPSVPDSASPADDSFVDPGERLYDLNMPAYVKFNYMAEREDELSLIKGTKVIVMEKCSDGWWRGSYNGQVGWFHSNYVTEEGDSPLGDNVGSLSEKLAAVVNNLNTGQVLHVVQALYPFSSSNDEELNFEKGDVMDVIEKPENDPEWWKCRKINGMVGLVPKNYVTVMQNNPLTSGLEPSPQQCDYIRPSLTGKFAGNPWYYGKVTRHQAEMALNERGHEGDFLIRDSESSPNDFSVSLKAQGKNKHFKVQLKETVYCIGQRKFSTMEELVEHYKKAPIFTSEQGEKLYLVKHLS | Full-length (residues 1–377) SH3 domains—underlined; SH2 domain—double underlined |
| SH3b(3) | MPAYVKFNYMAEREDELSLIKGTKVIVMEKSSDGWWRGSYNGQVGWFPSNYVTEEGDSPLSARKASIVKNLKDTLGIGKVKRKPSVPDSASPADDSFVDPGERLYDLNMPAYVKFNYMAEREDELSLIKGTKVIVMEKSSDGWWRGSYNGQVGWFPSNYVTEEGDSPLSARKASIVKNLKDTLGIGKVKRKPSVPDSASPADDSFVDPGERLYDLNMPAYVKFNYMAEREDELSLIKGTKVIVMEKSSDGWWRGSYNGQVGWFPSNYVTEEGDSPLNNPLTSGLEPSPPQSDYIRPSLTGKFAGNPWYYGKVTRHQAEMALNERGHEGDFLIRDSESSPNDFSVSLKAQGKNKHFKVQLKETVYSIGQRKFSTMEELVEHYKKAPIFTSEQGEKLYLVKHLS | N-terminal Nck1 region (residues 1–251) was replaced by three repeated SH3b (residues 109–168, underlined) domains tethered by Linker domain between SH3a and SH3b (residues 62–108). |
| SH3b(2) | MPAYVKFNYMAEREDELSLIKGTKVIVMEKSSDGWWRGSYNGQVGWFPSNYVTEEGDSPLSARKASIVKNLKDTLGIGKVKRKPSVPDSASPADDSFVDPGERLYDLNMPAYVKFNYMAEREDELSLIKGTKVIVMEKSSDGWWRGSYNGQVGWFPSNYVTEEGDSPLNNPLTSGLEPSPPQSDYIRPSLTGKFAGNPWYYGKVTRHQAEMALNERGHEGDFLIRDSESSPNDFSVSLKAQGKNKHFKVQLKETVYSIGQRKFSTMEELVEHYKKAPIFTSEQGEKLYLVKHLS | N-terminal Nck1 region (residues 1–251) was replaced by two repeated SH3b (residues 109–168, underlined) domains tethered by Linker domain between SH3a and SH3b (residues 61–108). |
| SH3b(1) | MPAYVKFNYMAEREDELSLIKGTKVIVMEKSSDGWWRGSYNGQVGWFPSNYVTEEGDSPLNNPLTSGLEPSPPQSDYIRPSLTGKFAGNPWYYGKVTRHQAEMALNERGHEGDFLIRDSESSPNDFSVSLKAQGKNKHFKVQLKETVYSIGQRKFSTMEELVEHYKKAPIFTSEQGEKLYLVKHLS | N-terminal Nck1 region (residues 1–251) was replaced by SH3b (residues 109–168, underlined) domain. |
| Src | WEIPRESLRLEVKLGQGCFGEVWMGTWNGTTRVAIKTLKPGTMSPEAFLQEAQVMKKLRHEKLVQLYAVVSEEPIYIVTEYMSKGSLLDFLKGETGKYLRLPQLVDMAAQIASGMAYVERMNYVHRDLRAANILVGENLVCKVADFGLARLIEDNEYTARQGAKFPIKWTAPEAALYGRFTIKSDVWSFGILLTELTTKGRVPYPGMVNREVLDQVERGYRMPCPPECP ESLHDLMCQCWRKEPEERPTFEYLQAFLEDY |
Kinase domain (residues 263–522) |
| FKBP | VQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNKPFKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVELLKLE | FKBP12 (residues 3–108) |
| FRB | ELIRVAILWHEMWHEGLEEASRLYFGERNVKGMFEVLEPLHAMMERGPQTLKETSFNQAYGRDLMEAQEWCRKYMKSGNVKDLTQAWDLYYHVFRRISKQ | Human mTOR (residues 2015–2114) |