IntAct European Bioinformatics Institute IntAct intact http://www.ebi.ac.uk/ http://www.ebi.ac.uk/intact/query/${ac} EBI-[0-9]+|IA:[0-9]+ INTerAction database (IntAct) provides an open source database and toolkit for the storage, presentation and analysis of molecular interactions. http://www.ebi.ac.uk/intact European Bioinformatics Institute; Wellcome Trust Genome Campus; Hinxton, Cambridge; CB10 1SD; United Kingdom http://www.ebi.ac.uk/intact/ Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. J. Biol. Chem. (0021-9258) 1999 imex curation Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. kauvarLM@earthlink.net 26-APR-2007: Contacted by JYOTI. Only protein-protein interactions human-t-cell Homo sapiens t-cell Peripheral blood T-lymphocytes anti bait coip anti bait coimmunoprecipitation anti bait coip western blot western blot Immuno blot Immuno blot Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. kauvarLM@earthlink.net Accepted 2007-MAY-10 by JYOTI. 26-APR-2007: Contacted by JYOTI. J. Biol. Chem. (0021-9258) 1999 Only protein-protein interactions imex curation Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. J. Biol. Chem. (0021-9258) 1999 imex curation Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. kauvarLM@earthlink.net 26-APR-2007: Contacted by JYOTI. Only protein-protein interactions yeasx Saccharomyces cerevisiae (Baker's yeast) Baker's yeast 2 hybrid two hybrid 2H 2h Gal4 transcription regeneration two-hybrid yeast two hybrid 2-hybrid classical two hybrid classical two hybrid 2H two-hybrid 2-hybrid Gal4 transcription regeneration 2h yeast two hybrid Y2H Y-2H 2 hybrid nucleotide sequence nucleotide sequence identification sequence cloning sequence cloning nucleotide sequence Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. kauvarLM@earthlink.net Accepted 2007-MAY-10 by JYOTI. 26-APR-2007: Contacted by JYOTI. Human T-cell cDNA J. Biol. Chem. (0021-9258) 1999 Only protein-protein interactions imex curation Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. J. Biol. Chem. (0021-9258) 1999 imex curation Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. kauvarLM@earthlink.net 26-APR-2007: Contacted by JYOTI. Only protein-protein interactions in vitro In vitro protein kinase assay protein kinase assay predetermined predetermined participant predetermined Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. kauvarLM@earthlink.net Accepted 2007-MAY-10 by JYOTI. 26-APR-2007: Contacted by JYOTI. J. Biol. Chem. (0021-9258) 1999 Only protein-protein interactions imex curation Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. J. Biol. Chem. (0021-9258) 1999 imex curation Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. kauvarLM@earthlink.net 26-APR-2007: Contacted by JYOTI. Only protein-protein interactions yeasx Saccharomyces cerevisiae (Baker's yeast) Baker's yeast 2 hybrid two hybrid 2H 2h Gal4 transcription regeneration two-hybrid yeast two hybrid 2-hybrid classical two hybrid classical two hybrid 2H two-hybrid 2-hybrid Gal4 transcription regeneration 2h yeast two hybrid Y2H Y-2H 2 hybrid predetermined predetermined participant predetermined Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. kauvarLM@earthlink.net Accepted 2007-MAY-10 by JYOTI. 26-APR-2007: Contacted by JYOTI. J. Biol. Chem. (0021-9258) 1999 Only protein-protein interactions imex curation Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. J. Biol. Chem. (0021-9258) 1999 imex curation Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. kauvarLM@earthlink.net 26-APR-2007: Contacted by JYOTI. Only protein-protein interactions in vitro In vitro elisa enzyme linked immunosorbent assay ELISA ELISA elisa antibody detection identification by antibody antibody detection Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. Polystyrene 96-well plates (Corning) were coated with indicated amounts of purified MBP-p59fyn. Recombinant, theta PKC or V1 fragments of theta PKC or delta PKC were added followed by incubation with peroxidase-conjugated anti-theta PKC antibody. Binding identified by development using the 3,3',5,5'-tetramethylbenzidine peroxidase substrate system kauvarLM@earthlink.net Accepted 2007-MAY-10 by JYOTI. 26-APR-2007: Contacted by JYOTI. J. Biol. Chem. (0021-9258) 1999 Only protein-protein interactions imex curation Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. Direct interaction in T-cells between thetaPKC and the tyrosine kinase p59fyn. J. Biol. Chem. (0021-9258) 1999 imex curation Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. kauvarLM@earthlink.net 26-APR-2007: Contacted by JYOTI. Only protein-protein interactions in vitro In vitro pull down pull down western blot western blot Immuno blot Immuno blot Ron D., Napolitano EW., Voronova A., Vasquez NJ., Roberts DN., Calio BL., Caothien RH., Pettiford SM., Wellik S., Mandac JB., Kauvar LM. kauvarLM@earthlink.net Accepted 2007-MAY-10 by JYOTI. 26-APR-2007: Contacted by JYOTI. J. Biol. Chem. (0021-9258) 1999 Only protein-protein interactions imex curation fyn_human Tyrosine-protein kinase Fyn p59-Fyn Proto-oncogene c-Fyn Src-like kinase FYN Proto-oncogene Syn protein protein human Homo sapiens Human MGCVQCKDKEATKLTEERDGSLNQSSGYRYGTDPTPQHYPSFGVTSIPNYNNFHAAGGQGLTVFGGVNSSSHTGTLRTRGGTGVTLFVALYDYEARTEDDLSFHKGEKFQILNSSEGDWWEARSLTTGETGYIPSNYVAPVDSIQAEEWYFGKLGRKDAERQLLSFGNPRGTFLIRESETTKGAYSLSIRDWDDMKGDHVKHYKIRKLDNGGYYITTRAQFETLQQLVQHYSERAAGLCCRLVVPCHKGMPRLTDLSVKTKDVWEIPRESLQLIKRLGNGQFGEVWMGTWNGNTKVAIKTLKPGTMSPESFLEEAQIMKKLKHDKLVQLYAVVSEEPIYIVTEYMNKGSLLDFLKDGEGRALKLPNLVDMAAQVAAGMAYIERMNYIHRDLRSANILVGNGLICKIADFGLARLIEDNEYTARQGAKFPIKWTAPEAALYGRFTIKSDVWSFGILLTELVTKGRVPYPGMNNREVLEQVERGYRMPCPQDCPISLHELMIHCWKKDPEERPTFEYLQSFLEDYFTATEPQYQPGENL DIP protein P06241 original sequence version: 195 DIP protein P06241 original sequence version: 147 DIP protein P06241 original sequence version: 164 4A1E443A4B5A0977 kpct_human Protein kinase C theta type PRKCQ nPKC-theta PRKCT protein protein human Homo sapiens Human MSPFLRIGLSNFDCGSCQSCQGEAVNPYCAVLVKEYVESENGQMYIQKKPTMYPPWDSTFDAHINKGRVMQIIVKGKNVDLISETTVELYSLAERCRKNNGKTEIWLELKPQGRMLMNARYFLEMSDTKDMNEFETEGFFALHQRRGAIKQAKVHHVKCHEFTATFFPQPTFCSVCHEFVWGLNKQGYQCRQCNAAIHKKCIDKVIAKCTGSAINSRETMFHKERFKIDMPHRFKVYNYKSPTFCEHCGTLLWGLARQGLKCDACGMNVHHRCQTKVANLCGINQKLMAEALAMIESTQQARCLRDTEQIFREGPVEIGLPCSIKNEARPPCLPTPGKREPQGISWESPLDEVDKMCHLPEPELNKERPSLQIKLKIEDFILHKMLGKGSFGKVFLAEFKKTNQFFAIKALKKDVVLMDDDVECTMVEKRVLSLAWEHPFLTHMFCTFQTKENLFFVMEYLNGGDLMYHIQSCHKFDLSRATFYAAEIILGLQFLHSKGIVYRDLKLDNILLDKDGHIKIADFGMCKENMLGDAKTNTFCGTPDYIAPEILLGQKYNHSVDWWSFGVLLYEMLIGQSPFHGQDEEELFHSIRMDNPFYPRWLEKEAKDLLVKLFVREPEKRLGVRGDIRQHPLFREINWEELERKEIDPPFRPKVKSPFDCSNFDKEFLNEKPRLSFADRALINSMDQNMFRNFSFMNPGMERLIS B3C53AB892D5210A kpcd_human Protein kinase C delta type nPKC-delta PRKCD Tyrosine-protein kinase PRKCD PKCD protein protein human Homo sapiens Human MAPFLRIAFNSYELGSLQAEDEANQPFCAVKMKEALSTERGKTLVQKKPTMYPEWKSTFDAHIYEGRVIQIVLMRAAEEPVSEVTVGVSVLAERCKKNNGKAEFWLDLQPQAKVLMSVQYFLEDVDCKQSMRSEDEAKFPTMNRRGAIKQAKIHYIKNHEFIATFFGQPTFCSVCKDFVWGLNKQGYKCRQCNAAIHKKCIDKIIGRCTGTAANSRDTIFQKERFNIDMPHRFKVHNYMSPTFCDHCGSLLWGLVKQGLKCEDCGMNVHHKCREKVANLCGINQKLLAEALNQVTQRASRRSDSASSEPVGIYQGFEKKTGVAGEDMQDNSGTYGKIWEGSSKCNINNFIFHKVLGKGSFGKVLLGELKGRGEYFAIKALKKDVVLIDDDVECTMVEKRVLTLAAENPFLTHLICTFQTKDHLFFVMEFLNGGDLMYHIQDKGRFELYRATFYAAEIMCGLQFLHSKGIIYRDLKLDNVLLDRDGHIKIADFGMCKENIFGESRASTFCGTPDYIAPEILQGLKYTFSVDWWSFGVLLYEMLIGQSPFHGDDEDELFESIRVDTPHYPRWITKESKDILEKLFEREPTKRLGVTGNIKIHPFFKTINWTLLEKRRLEPPFRPKVKSPRDYSNFDQEFLNEKARLSYSDKNLIDSMDQSAFAGFSFVNPKFEHLLED DIP protein Q05655 original sequence version: 92 DIP protein Q05655 original sequence version: 171 013F4314A2EE331A fyn-prkcq-1 6 7 unspecified role unspecified role prey prey human-jurkat Homo sapiens jurkat Human T cell leukemia 8 unspecified role unspecified role bait bait region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined region sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) physical association physical association Binding of p59fyn to theta PKC-V1 affinity sorbent Fig. 1 fyn-prkcq-2 5 8 unspecified role unspecified role prey prey n-terminal region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range region sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) 7 unspecified role unspecified role bait bait n-terminal mbp tag maltose binding protein tag maltose binding protein tag maltose binding protein tag mbp tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range physical association physical association To further establish that theta PKC can bind to p59fyn, purified recombinant constructs were used in a 96-well microplate binding assay (Fig. 2) Fig. 2 fyn-prkcd-1 5 7 unspecified role unspecified role bait bait n-terminal mbp tag maltose binding protein tag maltose binding protein tag maltose binding protein tag mbp tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range 9 unspecified role unspecified role prey prey region his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range c2 region sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) physical association physical association B, the V1 variable domains of theta PKC and delta PKC were compared for ability to bind MBP-p59fyn. Fig. 2 Reaction is weaker than that observed with PKC theta (PRKCQ) prkcq-2 3 8 putative self putative self putative self putative self n-terminal his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range spofr-sf_9 Spodoptera frugiperda insect cells Fall armyworm sf_9 insect cells Derived from pupal ovarian tissue of the fall armyworm Cloned from the parent cell line, IPLB-SF-21-AE (SF-21) derived from immature ovaries of fall armyworm Spodoptera frugiperda pupae cells are susceptible to Baculovirus infection phosphorylation phosphorylation reaction phosphorylation true the presence of p59fyn potentiated the kinase activity of theta PKC, increasing maximal phosphorylation of histone IIIs in an in vitro kinase assay (Fig. 3 lanes 7 and 8), with a more modest effect on autophosphorylation (lanes 3 and 4). 3 FYN (P06241) fyn-prkcq-3 1 8 unspecified role unspecified role bait bait 7 unspecified role unspecified role prey prey physical association physical association Co-immunoprecipitation from T-cells of theta PKC and p59fyn. Fig. 4 fyn-prkcq-4 1 8 unspecified role unspecified role prey prey 7 unspecified role unspecified role bait bait physical association physical association Co-immunoprecipitation from T-cells of theta PKC and p59fyn. Fig. 4 fyn-prkcq-5 3 7 enzyme enzyme unspecified role unspecified role region mbp tag maltose binding protein tag maltose binding protein tag maltose binding protein tag mbp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 8 enzyme target enzyme target substrate substrate unspecified role unspecified role n-terminal his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range v1 region sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) phosphorylation phosphorylation reaction phosphorylation In vitro phosphorylation of theta PKC-V1 by p59fyn. Fig. 5 fyn-prkcq-6 4 7 unspecified role unspecified role prey prey kinase region sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain P06241:p.Lys299Ala mutation decreasing mutation decreasing interaction hotspot hotspot mutation decreasing mutation analysis mutation analysis certain certain sequence position certain certain certain sequence position certain K A Wed May 02 14:45:23 BST 2018 This feature has been corrected as a result of our quality control procedures. The original label was 'lys299ala' Tue Jun 30 15:51:21 BST 2020 This feature has been corrected as a result of our quality control procedures. The original label was 'p.Lys299Ala' 8 unspecified role unspecified role bait bait v1 region sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain physical association physical association Co-immunoprecipitation from T-cells of theta PKC and p59fyn. Fig. 4 60uM TER14687, (+/-)-2-(N,N-dimethylaminomethyl)-1-indanone HCl, an inhibitor of IL-2 production in Jurkat T-cells Coordinaes of kinase domain taken from UniProtKB entry fyn-prkcd-2 3 7 enzyme enzyme unspecified role unspecified role region mbp tag maltose binding protein tag maltose binding protein tag maltose binding protein tag mbp tag undetermined undetermined sequence position undetermined undetermined undetermined sequence position undetermined 9 enzyme target enzyme target substrate substrate unspecified role unspecified role n-terminal his tag his tag Hexa-His-tag 6-His-tag Histidine-tag Hexa-His-tag Histidine-tag 6-His-tag n-term range n-terminal range n-term range n-term range n-terminal range n-term range v1 region sufficient to bind sufficient binding region sufficient to bind deletion analysis deletion analysis certain certain sequence position certain certain certain sequence position certain ecoli Escherichia coli (strain K12) phosphorylation phosphorylation reaction phosphorylation In vitro phosphorylation of theta PKC-V1 by p59fyn. Fig. 5 fyn-prkcq-7 2 8 unspecified role unspecified role bait bait 7 unspecified role unspecified role prey prey physical association physical association Likewise, screening a human cDNA library yielded several p59fyn clones but no p56lck clones. Interaction also proven using a mouse T-cell cDNA library spiked with human FYN but no indication as to source of prey isolated.