Mutational analysis of the Src SH3 domain: the same residues of the ligand binding surface are important for intra- and intermolecular interactions.

Erpel, Thorsten; Superti‐Furga, Giulio; Courtneidge, Sara A.

doi:10.1002/j.1460-2075.1995.tb07077.x

Cited by 154 publications

(175 citation statements)

References 75 publications

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“…Perturbations in the SH3 domain of other proteins were also shown to a ect their transforming potential (Erpel et al, 1995;Hirai and Varmus, 1990a,b;Jackson and Baltimore, 1989;Jackson et al, 1993;Kato et al, 1986;Mayer and Hanafusa, 1990;Seidel-Dugan et al, 1992;Weng et al, 1995). In these cases the mutations/ deletions were shown to a ect the biochemical properties such as enzymatic activity of the mutant proteins (Erpel et al, 1995;Hirai and Varmus, 1990a,b;Mayer and Hanafusa, 1990;Seidel-Dugan et al, 1992).…”

Section: Discussionmentioning

confidence: 99%

“…In these cases the mutations/ deletions were shown to a ect the biochemical properties such as enzymatic activity of the mutant proteins (Erpel et al, 1995;Hirai and Varmus, 1990a,b;Mayer and Hanafusa, 1990;Seidel-Dugan et al, 1992). We therefore wished to examine whether the biochemical properties of the mutant SH3 proteins di er from those of wild type Vav.…”

Section: Discussionmentioning

confidence: 99%

“…Mutations/deletions in the SH3 domain of various proteins have been shown to a ect their ability to associate with other proteins (Bustelo et al, 1995;Clark et al, 1992;Erpel et al, 1995;Hirai and Varmus, 1990a,b;Hobert et al, 1994;1996a,b;Jackson and Baltimore, 1989;Jackson et al, 1993;Kato et al, 1986;Mayer and Hanafusa, 1990;Romero et al, 1996 The immunoprecipitates were then incubated with lysates of cells overexpressing EGFR (HER14) that were either non induced (7) or induced (+) with 0.5 mg/ml EGF for 2 min. The proteins were then resolved on SDS ± PAGE and immunoblotted with anti-Ptyr Abs.…”

Section: Association Of Intact and Mutant Vavsh3c With Other Proteinsmentioning

confidence: 99%

“…The kinase activity of the SH3 mutant Src proteins increased relative to the intact protein and in addition, the cytoskeleton organization in cells with these proteins changed and resembled the morphology of v-src transformed cells (Erpel et al, 1995;Hirai and Varmus, 1990a,b;Kato et al, 1986;Seidel-Dugan et al, 1992;Weng et al, 1995). Detailed studies indicated that subtle amino-acid substitutions in the SH3 domain of Src changed their binding properties and therefore resulted in alteration of its biological functions (Erpel et al, 1995;Hirai and Varmus, 1990a,b;Kato et al, 1986;Seidel-Dugan et al, 1992;Weng et al, 1995).…”

Section: Introductionmentioning

confidence: 97%

“…Thus, mutations in the SH3 domain of the adaptor v-Crk (Mayer and Hanafusa, 1990) reduced its transforming potential whereas deletion/mutations of this domain from the proto-oncogenes Abl (Jackson and Baltimore, 1989;Jackson et al, 1993) and Src (Erpel et al, 1995;Hirai and Varmus, 1990a,b;Kato et al, 1986;Weng et al, 1995) increased their transforming potential. The kinase activity of the SH3 mutant Src proteins increased relative to the intact protein and in addition, the cytoskeleton organization in cells with these proteins changed and resembled the morphology of v-src transformed cells (Erpel et al, 1995;Hirai and Varmus, 1990a,b;Kato et al, 1986;Seidel-Dugan et al, 1992;Weng et al, 1995). Detailed studies indicated that subtle amino-acid substitutions in the SH3 domain of Src changed their binding properties and therefore resulted in alteration of its biological functions (Erpel et al, 1995;Hirai and Varmus, 1990a,b;Kato et al, 1986;Seidel-Dugan et al, 1992;Weng et al, 1995).…”

Section: Introductionmentioning

confidence: 99%

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Mutagenic analysis of Vav reveals that an intact SH3 domain is required for transformation

et al. 1998

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The vav proto-oncogene encodes a protein with multiple modulae domains that enable it to function as a mediator, linking tyrosine signaling to downstream events in hematopoietic cells. Circumstantial evidence suggests that protein-protein interactions exerted by two of these domains, the Src homology 2 (SH2) and the Src homology 3 (SH3), play an important role in the regulation of Vav activity. To study the relevance of the SH3 domain for the function of vav as a transforming gene, we have created several mutations in the SH3 domain located at its carboxy region. Substitution of the non-conserved aspartic acid 797 (to asparagine, D797N) retained the transforming potential of the vav oncogene, whereas substitutions of ®ve highly conserved amino-acids: alanine 789 (to asparagine, A789N), leucine 801 (to arginine, L801R), tryptophan 821 (to arginine, W821R), glycine 830 (to valine, G830V) and valine 837 (to glutamic acid, V837E) greatly reduced its transforming potential. The mutant proteins resemble Vav in many biochemical properties; however, while the transforming mutant protein (D797N) associates with several unidenti®ed proteins in a manner similar to that of Vav, the non-transforming mutant Vav proteins react very poorly with these proteins. Among the known Vav-interacting proteins, hnRNP-K associates with all mutant proteins except A789N and V837E whereas binding of Zyxin to any of the mutant proteins is not a ected. Taken together, our results clearly demonstrate that the SH3 domain has a positive e ect on vav activity and is needed for vav transformation. The vavSH3C associating protein(s) that are crucial for its activity as a transforming gene have probably not yet been identi®ed.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Association Of Intact and Mutant Vavsh3c With Other Proteinsmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 97%

Section: Introductionmentioning

confidence: 99%

See 3 more Smart Citations

Mutagenic analysis of Vav reveals that an intact SH3 domain is required for transformation

et al. 1998

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Disruption of a putative SH3 domain and the proline-rich motifs in the 53-kDa substrate of the insulin receptor kinase does not alter its subcellular localization or ability to serve as a substrate

Yeh

Li²,

Keller

et al. 1998

J. Cell. Biochem.

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The recently identified 53-kDa substrate of the insulin receptor family was further characterized in several retroviral-generated stable cell lines overexpressing the wild type and various mutant forms of the protein. To facilitate the study of its subcellular localization in NIH3T3 cells overexpressing insulin receptor, a myc epitope-tag was added to the carboxy terminus of the 53-kDa protein. Like the endogenous protein in Chinese hamster ovary cells, the expressed myc-tagged 53-kDa protein was found partially in the particulate fraction and was tyrosine phosphorylated in insulin-stimulated cells. Immunofluorescence studies showed for the first time that a fraction of the 53-kDa protein was localized to the plasma membrane. Confocal microscopy of cells double-labeled with antibodies to the insulin receptor and the myc epitope showed the two proteins co-localize at the plasma membrane at the level of light microscopy. Further analyses of the protein sequence of the 53-kDa substrate revealed the presence of a putative SH3 domain and two proline-rich regions, putative binding sites for SH3 and WW domains. Disruption of these three motifs by the introduction of previously characterized point mutations did not affect the membrane localization of the 53-kDa protein, its ability to serve as substrate of the insulin receptor, or its colocalization with the insulin receptor, suggesting these domains are not important in the subcellular targeting of the protein and instead may function in the interaction with subsequent signaling proteins.

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The SH3 domain of Bruton's tyrosine kinase displays altered ligand binding properties when auto‐phosphorylatedin vitro

et al. 1999

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Btk is a member of the Tec family of protein tyrosine kinases expressed in B cells. It is stimulated following cross‐linking of the B cell receptor which leads to the autophosphorylation of a specific residue in the SH3 domain, Y223. Previous work using Btk‐derived fusion proteins has shown that the Btk SH3 domain binds to c‐Cbl and Wiskott‐Aldrich syndrome protein (WASP) in vitro. We show here that when the Btk SH3 domain fusion protein is autophosphorylated, its ability to take part in protein interactions is altered as compared to the non‐phosphorylated fusion protein. Although the phosphorylated Btk SH3 domain still binds c‐Cbl, it no longer binds WASP and instead acquires a high affinity for kinase‐active Syk. The region of Syk responsible for this interaction is contained within its C terminus, suggesting a novel mechanism by which these proteins may interact.

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Mutational analysis of the Src SH3 domain: the same residues of the ligand binding surface are important for intra- and intermolecular interactions.

Cited by 154 publications

References 75 publications

Mutagenic analysis of Vav reveals that an intact SH3 domain is required for transformation

Mutagenic analysis of Vav reveals that an intact SH3 domain is required for transformation

Disruption of a putative SH3 domain and the proline-rich motifs in the 53-kDa substrate of the insulin receptor kinase does not alter its subcellular localization or ability to serve as a substrate

The SH3 domain of Bruton's tyrosine kinase displays altered ligand binding properties when auto‐phosphorylatedin vitro

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