Probing the Native Structure of Stathmin and Its Interaction Domains with Tubulin

Stathmin is a cytosoluble phosphoprotein proposed to be a regulatory relay integrating diverse intracellular signaling pathway. Its interaction with tubulin modulates microtubule dynamics by destabilization of assembled microtubules or inhibition of their polymerization from free tubulin. The aim of this study was to probe the native structure of stathmin and to delineate its minimal region able to interact with tubulin. Limited proteolysis of stathmin revealed four structured domains within the native protein, corresponding to amino acid sequences 22–81 (I), 95–113 (II), 113–128 (III), and 128–149 (IV), which allows us to propose stathmin folding hypotheses. Furthermore, stathmin proteolytic fragments were mixed to interact with tubulin, and those that retained affinity for tubulin were isolated by size exclusion chromatography and identified by matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. The results indicate that, to interact with tubulin, a stathmin fragment must span a minimal core region from residues 42 to 126, which interestingly corresponds to the predicted α-helical “interaction region” of stathmin. In addition, an interacting stathmin fragment must include a short N- or C-terminal extension. The functional significance of these interaction constrains is further validated by tubulin polymerization inhibition assays with fragments designed on the basis of the tubulin binding results. The present results will help to optimize further stathmin structural studies and to develop molecular tools to target its interaction with tubulin.

• Publication year

  2000

• Venue

  Journal of Biological Chemistry

• Publication date

  2000-03-10

• Fields of study

  Biology, Medicine, Chemistry

• Identifiers
  DOI 10.1074/jbc.275.10.6841PMID 10702243
• External record

  Open on Semantic Scholar

• Source metadata

  Semantic Scholar, PubMed

• No claims are published for this paper.

• No concepts are published for this paper.

• Molecular Cloning: A Laboratory Manual

  2001cited by this paper
• Op18/Stathmin Mediates Multiple Region-Specific Tubulin and Microtubule-Regulating Activities

  1999cited by this paper
• Control of K+ channel gating by protein phosphorylation: structural switches of the inactivation gate

  1999cited by this paper
• Stathmin interaction with HSC70 family proteins

  1999cited by this paper
• Dissociation of the tubulin-sequestering and microtubule catastrophe-promoting activities of oncoprotein 18/stathmin.

  1999cited by this paper
• The stathmin phosphoprotein family: intracellular localization and effects on the microtubule network.

  1998cited by this paper
• Retrograde Transport of Golgi-localized Proteins to the ER

  1998cited by this paper
• Identification of in Vitro Phosphorylation Sites in the Growth Cone Protein SCG10

  1998cited by this paper
• Regulation of microtubule dynamics by Ca2+/calmodulin-dependent kinase IV/Gr-dependent phosphorylation of oncoprotein 18

  1997cited by this paper
• The stathmin family -- molecular and biological characterization of novel mammalian proteins expressed in the nervous system.

  1997cited by this paper
• KIS Is a Protein Kinase with an RNA Recognition Motif*

  1997cited by this paper
• Exploring the limits of nearest neighbour secondary structure prediction.

  1997cited by this paper
• The Microtubule-destabilizing Activity of Metablastin (p19) Is Controlled by Phosphorylation*

  1997cited by this paper
• Stathmin: a tubulin-sequestering protein which forms a ternary T2S complex with two tubulin molecules.

  1997cited by this paper
• Probing protein structure using biochemical and biophysical methods. Proteolysis, matrix-assisted laser desorption/ionization mass spectrometry, high-performance liquid chromatography and size-exclusion chromatography of p21Waf1/Cip1/Sdi1.

  1997cited by this paper
• Regulation of microtubule dynamics by the neuronal growth-associated protein SCG10.

  1997cited by this paper
• Phosphorylation regulates the microtubule‐destabilizing activity of stathmin and its interaction with tubulin

  1997cited by this paper
• The TSG101 tumor susceptibility gene is located in chromosome 11 band p15 and is mutated in human breast cancer.

  1997cited by this paper
• Identification of a protein that interacts with tubulin dimers and increases the catastrophe rate of microtubules.

  1996cited by this paper
• Oncoprotein 18 is a phosphorylation‐responsive regulator of microtubule dynamics.

  1996cited by this paper
• Tsg101: a novel tumor susceptibility gene isolated by controlled homozygous functional knockout of allelic loci in mammalian cells.

  1996cited by this paper
• Stathmin interaction with a putative kinase and coiled-coil-forming protein domains.

  1995cited by this paper
• Probing the solution structure of the DNA‐binding protein Max by a combination of proteolysis and mass spectrometry

  1995cited by this paper
• SOPMA: significant improvements in protein secondary structure prediction by consensus prediction from multiple alignments

  1995cited by this paper
• Modeling studies of the change in conformation required for cleavage of limited proteolytic sites

  1994cited by this paper
• Molecular characterization of human stathmin expressed in Escherichia coli: site-directed mutagenesis of two phosphorylatable serines (Ser-25 and Ser-63).

  1994cited by this paper
• The phosphorylation of stathmin by MAP kinase

  1993cited by this paper
• Stathmin Phosphorylation Is Regulated in Striatal Neurons by Vasoactive Intestinal Peptide and Monoamines via Multiple Intracellular Pathways

  1992cited by this paper
• Glycogen phosphorylase: control by phosphorylation and allosteric effectors

  1992cited by this paper
• Stathmin: a relay phosphoprotein for multiple signal transduction?

  1991cited by this paper
• Characterization of the gene for a proliferation-related phosphoprotein (oncoprotein 18) expressed in high amounts in acute leukemia.

  1991cited by this paper
• A single amino acid difference distinguishes the human and the rat sequences of stathmin, a ubiquitous intracellular phosphoprotein associated with cell regulations

  1990cited by this paper
• The "megaprimer" method of site-directed mutagenesis.

  1990cited by this paper
• Developmental tissue expression and phylogenetic conservation of stathmin, a phosphoprotein associated with cell regulations.

  1990cited by this paper
• N-myc gene amplification in neuroblastoma is associated with altered phosphorylation of a proliferation related polypeptide (Op18).

  1990cited by this paper
• Phosphorylation of stathmin and other proteins related to nerve growth factor-induced regulation of PC12 cells.

  1990cited by this paper
• Intracellular substrates for extracellular signaling. Characterization of a ubiquitous, neuron-enriched phosphoprotein (stathmin).

  1989cited by this paper
• Phosphorylation of a group of proteins related to the physiological, multihormonal regulations of the various cell types in the anterior pituitary gland.

  1989cited by this paper
• Phosphorylation of intracellular proteins related to the multihormonal regulation of prolactin: comparison of normal anterior pituitary cells in culture with the tumor-derived GH cell lines.

  1988cited by this paper
• Properties of p19, a novel cAMP-dependent protein kinase substrate protein purified from bovine brain.

  1987cited by this paper
• Correlation between sites of limited proteolysis and segmental mobility in thermolysin.

  1986cited by this paper
• Dynamic instability of microtubule growth

  1984cited by this paper
• Distinct patterns of cytoplasmic protein phosphorylation related to regulation of synthesis and release of prolactin by GH cells.

  1983cited by this paper
• Analysis of the accuracy and implications of simple methods for predicting the secondary structure of globular proteins.

  1978cited by this paper
• Empirical predictions of protein conformation.

  1978cited by this paper

• Microtubules, Membranes, and Movement: New Roles for Stathmin‐2 in Axon Integrity

  2024cites this paper
• The Mechanism of Tubulin Assembly into Microtubules: Insights from Structural Studies

  2020cites this paper
• Alternation of Mitochondrial and Golgi Apparatus in Neurodegenerative Disorders

  2019cites this paper
• Somatic environment and germinal differentiation in antral follicle: The effect of FSH withdrawal and basal LH on oocyte competence acquisition in cattle.

  2016cites this paper
• Mass spectrometry based strategy for studies of binding sites and structural changes of cisplatin binding to myoglobin.

  2016cites this paper
• Neuronal stathmins: a family of phosphoproteins cooperating for neuronal development, plasticity and regeneration.

  2015cites this paper
• Combinatory use of cell-free protein expression, limited proteolysis and mass spectrometry for the high-throughput protein domain identification.

  2014cites this paper
• Feedback mechanism for microtubule length regulation by stathmin gradients.

  2014cites this paper
• A hypothesis on the origin and evolution of tubulin.

  2013cites this paper
• Cytoskeleton and Human Disease

  2012cites this paper
• Stathmin and Cancer

  2012cites this paper
• Specific Serine-Proline Phosphorylation and Glycogen Synthase Kinase 3β-directed Subcellular Targeting of Stathmin 3/Sclip in Neurons*

  2012cites this paper
• Systematic Identification of Tubulin-interacting Fragments of the Microtubule-associated Protein Tau Leads to a Highly Efficient Promoter of Microtubule Assembly*

  2011cites this paper
• The binding of vinca domain agents to tubulin: structural and biochemical studies.

  2010cites this paper
• Aurora kinase inhibitor PHA-739358 suppresses growth of hepatocellular carcinoma in vitro and in a xenograft mouse model.

  2009cites this paper
• Caractérisation d’une nouvelle protéine associée aux microtubules, SL21

  2008cites this paper
• Stathmin 1: a novel therapeutic target for anticancer activity

  2008cites this paper
• Peptide P5 (residues 628–683), comprising the entire membrane proximal region of HIV-1 gp41 and its calcium-binding site, is a potent inhibitor of HIV-1 infection

  2008cites this paper
• Vibrational signatures of protonated, phosphorylated amino acids in the gas phase.

  2008cites this paper
• Insight into the GTPase activity of tubulin from complexes with stathmin-like domains.

  2007cites this paper
• Structure and thermodynamics of the tubulin-stathmin interaction.

  2007cites this paper
• The interplay between structure and function in intrinsically unstructured proteins

  2005cites this paper
• Etude des effets des peptides amyloïdes : du fonctionnement de la synapse aux modifications du cytosquelette dans l'apoptose neuronale.

  2005cites this paper
• Structural basis for the regulation of tubulin by vinblastine

  2005cites this paper
• N-terminal stathmin-like peptides bind tubulin and impede microtubule assembly.

  2005cites this paper
• Matrix-assisted laser desorption/ionization analysis of non-covalent complexes: fundamentals and applications.

  2005cites this paper
• Role of the lactoferrin-binding protein in pathogenesis of streptococcus uberis

  2005cites this paper
• Evaluation of cardosin A as a proteolytic probe in the presence of organic solvents

  2004cites this paper
• A synergistic relationship between three regions of stathmin family proteins is required for the formation of a stable complex with tubulin.

  2004cites this paper
• Thermodynamics of the Op18/Stathmin-Tubulin Interaction*

  2003cites this paper
• [The stathmin-tubulin interaction and the regulation of the microtubule assembly].

  2003cites this paper
• L’interaction stathmine–tubuline et la régulation de la dynamique des microtubules

  2003cites this paper
• Stathmin binds heat-shock protein 70

  2003cites this paper
• The functional benefits of protein disorder

  2003cites this paper
• The oncoprotein 18/stathmin family of microtubule destabilizers.

  2002cites this paper
• SCG10‐related neuronal growth‐associated proteins in neural development, plasticity, degeneration, and aging

  2002cites this paper
• Drosophila stathmin: a microtubule-destabilizing factor involved in nervous system formation.

  2002cites this paper
• Stathmin Family Proteins Display Specific Molecular and Tubulin Binding Properties*

  2001cites this paper
• Differential response of mature TrkA/p75NTR expressing human and pig oligodendrocytes: Aging, does it matter?

  2001cites this paper
• Aberrant mobility phenomena of the DNA repair protein XPA

  2001cites this paper
• Identification by mass spectrometry of two‐dimensional gel electrophoresis‐separated proteins extracted from lager brewing yeast

  2001cites this paper
• Localization of neuronal growth-associated, microtubule-destabilizing factor SCG10 in brain-derived raft membrane microdomains.

  2001cites this paper
• Isotope-tagged cross-linking reagents. A new tool in mass spectrometric protein interaction analysis.

  2001cites this paper
• The 4 A X-ray structure of a tubulin:stathmin-like domain complex.

  2000cites this paper
• Mutational Analysis of Op18/Stathmin-Tubulin-interacting Surfaces

  2000cites this paper