Structural basis for activation of human lymphocyte kinase Lck upon tyrosine phosphorylation

REGULATION through phosphorylation is a characteristic of signalling pathways1,2, and the lymphocyte kinase Lck (p56 lck) both performs phosphorylation and is affected by it. Lck is a Src-family tyrosine kinase expressed in T lymphocytes, where it participates in the cellular immune response 3. Like all Src homologues, it comprises SH3, SH2 and kinase domains. Lck associates through its distinctive ammo-terminal segment with the cytoplasmic tails of either T-cell co-receptor, CD4 or CD8-α 4,5. Activated Lck phosphorylates T-cell receptor ε-chains, which then recruit the ZAP70 kinase to promote T-cell activation 6. Lck is activated by autophosphorylation at Tyr394 in the activation loop 7 and it is inactive when Tyr 505 near the carboxy terminus is phosphorylated and interacts with its own SH2 domain8. Here we report the crystal structure of the Lck tyrosine kinase domain (LCKK) in its activated state at 1.7 Å resolution. The structure reveals how a phosphoryl group at Tyr 394 generates a competent active site. Comparisons with other kinase structures indicate that tyrosine phophophorylation and ligand binding may in general elicit two distinct hinge-like movements between the kinase subdomains. From modelling studies, we suggest a basis for inhibition by phosphorylation at Tyr 505.

[1]  Susan S. Taylor,et al.  Crystal structures of the myristylated catalytic subunit of cAMP‐dependent protein kinase reveal open and closed conformations , 1993, Protein science : a publication of the Protein Society.

[2]  Philip D. Jeffrey,et al.  Crystal structure of the p27Kip1 cyclin-dependent-kinase inibitor bound to the cyclin A–Cdk2 complex , 1996, Nature.

[3]  M. Fournel,et al.  The CD4 associated tyrosine protein kinase p56lck is positively regulated through its site of autophosphorylation. , 1990, Oncogene.

[4]  R. Perlmutter,et al.  Interaction of the unique N-terminal region of tyrosine kinase p56 lck with cytoplasmic domains of CD4 and CD8 is mediated by cysteine motifs , 1990, Cell.

[5]  J. Corbin,et al.  Structure and function of cyclic nucleotide-dependent protein kinases. , 1994, Annual review of physiology.

[6]  K. Titani,et al.  Sequence of two phosphorylated sites in the catalytic subunit of bovine cardiac muscle adenosine 3':5'-monophosphate-dependent protein kinase. , 1979, The Journal of biological chemistry.

[7]  A. Weiss,et al.  Sequential interactions of the TCR with two distinct cytoplasmic tyrosine kinases. , 1994, Science.

[8]  Joseph Schlessinger,et al.  Structure of the FGF Receptor Tyrosine Kinase Domain Reveals a Novel Autoinhibitory Mechanism , 1996, Cell.

[9]  Jonathan A. Cooper,et al.  The when and how of Src regulation , 1993, Cell.

[10]  R. Perlmutter,et al.  Structure and expression of Ick transcripts in human lymphoid cells , 1988 .

[11]  Axel T. Brunger,et al.  X-PLOR Version 3.1: A System for X-ray Crystallography and NMR , 1992 .

[12]  S. Hubbard,et al.  Crystal structure of the tyrosine kinase domain of the human insulin receptor , 1994, Nature.

[13]  Michael J. Eck,et al.  Structure of the regulatory domains of the Src-family tyrosine kinase Lck , 1994, Nature.

[14]  Hong Sun,et al.  The coordinated action of protein tyrosine phosphatases and kinases in cell signaling. , 1994, Trends in biochemical sciences.

[15]  S. Taylor,et al.  Domain movements in protein kinases. , 1994, Current opinion in structural biology.

[16]  D. Stern,et al.  The Ick tyrosine protein kinase interacts with the cytoplasmic tail of the CD4 glycoprotein through its unique amino-terminal domain , 1989, Cell.

[17]  K. Sharp,et al.  Protein folding and association: Insights from the interfacial and thermodynamic properties of hydrocarbons , 1991, Proteins.

[18]  E. Krebs,et al.  Conversion of phosphorylase b to phosphorylase a in muscle extracts. , 1955, The Journal of biological chemistry.

[19]  J. Zheng,et al.  Crystal structure of the catalytic subunit of cyclic adenosine monophosphate-dependent protein kinase. , 1991, Science.

[20]  L. Cantley,et al.  Oncogenes and signal transduction , 1991, Cell.

[21]  J. Zou,et al.  Improved methods for building protein models in electron density maps and the location of errors in these models. , 1991, Acta crystallographica. Section A, Foundations of crystallography.

[22]  T. Pawson,et al.  Regulation of the enzymatic function of the lymphocyte-specific tyrosine protein kinase p56lck by the non-catalytic SH2 and SH3 domains. , 1992, Oncogene.

[23]  P. Jeffrey,et al.  Structural basis of cyclin-dependent kinase activation by phosphorylation , 1996, Nature Structural Biology.

[24]  L. Johnson,et al.  Active and Inactive Protein Kinases: Structural Basis for Regulation , 1996, Cell.

[25]  W. Hendrickson.  Determination of macromolecular structures from anomalous diffraction of synchrotron radiation. , 1991, Science.

[26]  S V Evans,et al.  SETOR: hardware-lighted three-dimensional solid model representations of macromolecules. , 1993, Journal of molecular graphics.