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Title: Structural Features and Chaperone Activity of the NudC Protein Family

Abstract

The NudC family consists of four conserved proteins with representatives in all eukaryotes. The archetypal nudC gene from Aspergillus nidulans is a member of the nud gene family that is involved in the maintenance of nuclear migration. This family also includes nudF, whose human orthologue, Lis1, codes for a protein essential for brain cortex development. Three paralogues of NudC are known in vertebrates: NudC, NudC-like (NudCL), and NudC-like 2 (NudCL2). The fourth distantly related member of the family, CML66, contains a NudC-like domain. The three principal NudC proteins have no catalytic activity but appear to play as yet poorly defined roles in proliferating and dividing cells. We present crystallographic and NMR studies of the human NudC protein and discuss the results in the context of structures recently deposited by structural genomics centers (i.e., NudCL and mouse NudCL2). All proteins share the same core CS domain characteristic of proteins acting either as cochaperones of Hsp90 or as independent small heat shock proteins. However, while NudC and NudCL dimerize via an N-terminally located coiled coil, the smaller NudCL2 lacks this motif and instead dimerizes as a result of unique domain swapping. We show that NudC and NudCL, but not NudCL2, inhibit themore » aggregation of several target proteins, consistent with an Hsp90-independent heat shock protein function. Importantly, and in contrast to several previous reports, none of the three proteins is able to form binary complexes with Lis1. The availability of structural information will be of help in further studies on the cellular functions of the NudC family.« less

Authors:
; ; ; ; ; ; ; ; ; ; ; ;  [1];  [2]
  1. (UV)
  2. (
Publication Date:
Research Org.:
Argonne National Lab. (ANL), Argonne, IL (United States). Advanced Photon Source (APS)
Sponsoring Org.:
National Institutes of Health (NIH)
OSTI Identifier:
1040895
Resource Type:
Journal Article
Resource Relation:
Journal Name: Journal of Molecular Biology; Journal Volume: 409; Journal Issue: 5
Country of Publication:
United States
Language:
ENGLISH
Subject:
59 BASIC BIOLOGICAL SCIENCES; 60 APPLIED LIFE SCIENCES; ASPERGILLUS; AVAILABILITY; BRAIN; CRYSTALLOGRAPHY; GENES; HEAT-SHOCK PROTEINS; MAINTENANCE; PROTEIN STRUCTURE; PROTEINS; TARGETS; VERTEBRATES

Citation Formats

Zheng, Meiying, Cierpicki, Tomasz, Burdette, Alexander J., Utepbergenov, Darkhan, Janczyk, Pawe, #322, , #321, ., Derewenda, Urszula, Stukenberg, P. Todd, Caldwell, Kim A., Derewenda, Zygmunt S., and UAT). Structural Features and Chaperone Activity of the NudC Protein Family. United States: N. p., 2012. Web. doi:10.1016/j.jmb.2011.04.018.
Zheng, Meiying, Cierpicki, Tomasz, Burdette, Alexander J., Utepbergenov, Darkhan, Janczyk, Pawe, #322, , #321, ., Derewenda, Urszula, Stukenberg, P. Todd, Caldwell, Kim A., Derewenda, Zygmunt S., & UAT). Structural Features and Chaperone Activity of the NudC Protein Family. United States. doi:10.1016/j.jmb.2011.04.018.
Zheng, Meiying, Cierpicki, Tomasz, Burdette, Alexander J., Utepbergenov, Darkhan, Janczyk, Pawe, #322, , #321, ., Derewenda, Urszula, Stukenberg, P. Todd, Caldwell, Kim A., Derewenda, Zygmunt S., and UAT). 2012. "Structural Features and Chaperone Activity of the NudC Protein Family". United States. doi:10.1016/j.jmb.2011.04.018.
@article{osti_1040895,
title = {Structural Features and Chaperone Activity of the NudC Protein Family},
author = {Zheng, Meiying and Cierpicki, Tomasz and Burdette, Alexander J. and Utepbergenov, Darkhan and Janczyk, Pawe and #322 and and #321 and . and Derewenda, Urszula and Stukenberg, P. Todd and Caldwell, Kim A. and Derewenda, Zygmunt S. and UAT)},
abstractNote = {The NudC family consists of four conserved proteins with representatives in all eukaryotes. The archetypal nudC gene from Aspergillus nidulans is a member of the nud gene family that is involved in the maintenance of nuclear migration. This family also includes nudF, whose human orthologue, Lis1, codes for a protein essential for brain cortex development. Three paralogues of NudC are known in vertebrates: NudC, NudC-like (NudCL), and NudC-like 2 (NudCL2). The fourth distantly related member of the family, CML66, contains a NudC-like domain. The three principal NudC proteins have no catalytic activity but appear to play as yet poorly defined roles in proliferating and dividing cells. We present crystallographic and NMR studies of the human NudC protein and discuss the results in the context of structures recently deposited by structural genomics centers (i.e., NudCL and mouse NudCL2). All proteins share the same core CS domain characteristic of proteins acting either as cochaperones of Hsp90 or as independent small heat shock proteins. However, while NudC and NudCL dimerize via an N-terminally located coiled coil, the smaller NudCL2 lacks this motif and instead dimerizes as a result of unique domain swapping. We show that NudC and NudCL, but not NudCL2, inhibit the aggregation of several target proteins, consistent with an Hsp90-independent heat shock protein function. Importantly, and in contrast to several previous reports, none of the three proteins is able to form binary complexes with Lis1. The availability of structural information will be of help in further studies on the cellular functions of the NudC family.},
doi = {10.1016/j.jmb.2011.04.018},
journal = {Journal of Molecular Biology},
number = 5,
volume = 409,
place = {United States},
year = 2012,
month = 5
}
  • No abstract prepared.
  • No abstract prepared.
  • Protease activity assays are important for elucidating protease function and for developing new therapeutic agents. In this study, a novel turbidimetric method for determining the protease activity using a protease-responsive chaperone protein is described. For this purpose, a recombinant small heat-shock protein (sHSP) with an introduced Factor Xa protease recognition site was synthesized in bacteria. This recombinant mutant, FXa-HSP, exhibited chaperone-like activity at high temperatures in cell lysates. However, the chaperone-like activity of FXa-HSP decreased dramatically following treatment with Factor Xa. Protein precipitation was subsequently observed in the cell lysates. The reaction was Factor Xa concentration-dependent and was quantitatively suppressedmore » by a specific inhibitor for Factor Xa. Protein aggregation was detected by a simple method based on turbidimetry. The results clearly demonstrate that this assay is an effective, easy-to-use method for determining protease activities without the requirement of labeling procedures and the use of radioisotopes.« less
  • The plague-causing bacterium Yersinia pestis utilizes a type III secretion system to deliver effector proteins into mammalian cells where they interfere with signal transduction pathways that mediate phagocytosis and the inflammatory response. Effector proteins are injected through a hollow needle structure composed of the protein YscF. YscG and YscE act as 'chaperones' to prevent premature polymerization of YscF in the cytosol of the bacterium prior to assembly of the needle. Here, we report the crystal structure of the YscEFG protein complex at 1.8 {angstrom} resolution. Overall, the structure is similar to that of the analogous PscEFG complex from the Pseudomonasmore » aeruginosa type III secretion system, but there are noteworthy differences. The structure confirms that, like PscG, YscG is a member of the tetratricopeptide repeat family of proteins. YscG binds tightly to the C-terminal half of YscF, implying that it is this region of YscF that controls its polymerization into the needle structure. YscE interacts with the N-terminal tetratricopeptide repeat motif of YscG but makes very little direct contact with YscF. Its function may be to stabilize the structure of YscG and/or to participate in recruiting the complex to the secretion apparatus. No electron density could be observed for the 49 N-terminal residues of YscF. This and additional evidence suggest that the N-terminus of YscF is disordered in the complex with YscE and YscG. As expected, conserved residues in the C-terminal half of YscF mediate important intra- and intermolecular interactions in the complex. Moreover, the phenotypes of some previously characterized mutations in the C-terminal half of YscF can be rationalized in terms of the structure of the heterotrimeric YscEFG complex.« less