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Title: The Thioredoxin Domain of Neisseria Gonorrhoeae PilB can use Electrons from DsbD to Reduce Downstream Methionine Sulfoxide Reductases

Abstract

The PilB protein from Neisseria gonorrhoeae is located in the periplasm and made up of three domains. The N-terminal, thioredoxin-like domain (NT domain) is fused to tandem methionine sulfoxide reductase A and B domains (MsrA/B). We show that the {alpha} domain of Escherichia coli DsbD is able to reduce the oxidized NT domain, which suggests that DsbD in Neisseria can transfer electrons from the cytoplasmic thioredoxin to the periplasm for the reduction of the MsrA/B domains. An analysis of the available complete genomes provides further evidence for this proposition in other bacteria where DsbD/CcdA, Trx, MsrA, and MsrB gene homologs are all located in a gene cluster with a common transcriptional direction. An examination of wild-type PilB and a panel of Cys to Ser mutants of the full-length protein and the individually expressed domains have also shown that the NT domain more efficiently reduces the MsrA/B domains when in the polyprotein context. Within this framework there does not appear to be a preference for the NT domain to reduce the proximal MsrA domain over MsrB domain. Finally, we report the 1.6 {angstrom} crystal structure of the NT domain. This structure confirms the presence of a surface loop that makes itmore » different from other membrane-tethered, Trx-like molecules including TlpA, CcmG and ResA. Subtle differences are observed in this loop when compared to the N. meningitidis NT domain structure. The data taken together supports the formation of specific NT domain interactions with the MsrA/B domains and its in vivo recycling partner, DsbD.« less

Authors:
; ; ; ; ;
Publication Date:
Research Org.:
Brookhaven National Laboratory (BNL) National Synchrotron Light Source
Sponsoring Org.:
Doe - Office Of Science
OSTI Identifier:
930583
Report Number(s):
BNL-80796-2008-JA
Journal ID: ISSN 0021-9258; JBCHA3; TRN: US200904%%796
DOE Contract Number:
DE-AC02-98CH10886
Resource Type:
Journal Article
Resource Relation:
Journal Name: Journal of Biological Chemistry; Journal Volume: 281
Country of Publication:
United States
Language:
English
Subject:
36 MATERIALS SCIENCE; BACTERIA; CRYSTAL STRUCTURE; DOMAIN STRUCTURE; ELECTRONS; ESCHERICHIA COLI; GENES; IN VIVO; METHIONINE; MUTANTS; OXIDOREDUCTASES; PROTEINS; RECYCLING; SULFOXIDES; national synchrotron light source

Citation Formats

Brot,N., Collet, J., Johnson, L., Jonsson, T., Weissbach, H., and Lowther, W.. The Thioredoxin Domain of Neisseria Gonorrhoeae PilB can use Electrons from DsbD to Reduce Downstream Methionine Sulfoxide Reductases. United States: N. p., 2006. Web. doi:10.1074/jbc.M604971200.
Brot,N., Collet, J., Johnson, L., Jonsson, T., Weissbach, H., & Lowther, W.. The Thioredoxin Domain of Neisseria Gonorrhoeae PilB can use Electrons from DsbD to Reduce Downstream Methionine Sulfoxide Reductases. United States. doi:10.1074/jbc.M604971200.
Brot,N., Collet, J., Johnson, L., Jonsson, T., Weissbach, H., and Lowther, W.. Sun . "The Thioredoxin Domain of Neisseria Gonorrhoeae PilB can use Electrons from DsbD to Reduce Downstream Methionine Sulfoxide Reductases". United States. doi:10.1074/jbc.M604971200.
@article{osti_930583,
title = {The Thioredoxin Domain of Neisseria Gonorrhoeae PilB can use Electrons from DsbD to Reduce Downstream Methionine Sulfoxide Reductases},
author = {Brot,N. and Collet, J. and Johnson, L. and Jonsson, T. and Weissbach, H. and Lowther, W.},
abstractNote = {The PilB protein from Neisseria gonorrhoeae is located in the periplasm and made up of three domains. The N-terminal, thioredoxin-like domain (NT domain) is fused to tandem methionine sulfoxide reductase A and B domains (MsrA/B). We show that the {alpha} domain of Escherichia coli DsbD is able to reduce the oxidized NT domain, which suggests that DsbD in Neisseria can transfer electrons from the cytoplasmic thioredoxin to the periplasm for the reduction of the MsrA/B domains. An analysis of the available complete genomes provides further evidence for this proposition in other bacteria where DsbD/CcdA, Trx, MsrA, and MsrB gene homologs are all located in a gene cluster with a common transcriptional direction. An examination of wild-type PilB and a panel of Cys to Ser mutants of the full-length protein and the individually expressed domains have also shown that the NT domain more efficiently reduces the MsrA/B domains when in the polyprotein context. Within this framework there does not appear to be a preference for the NT domain to reduce the proximal MsrA domain over MsrB domain. Finally, we report the 1.6 {angstrom} crystal structure of the NT domain. This structure confirms the presence of a surface loop that makes it different from other membrane-tethered, Trx-like molecules including TlpA, CcmG and ResA. Subtle differences are observed in this loop when compared to the N. meningitidis NT domain structure. The data taken together supports the formation of specific NT domain interactions with the MsrA/B domains and its in vivo recycling partner, DsbD.},
doi = {10.1074/jbc.M604971200},
journal = {Journal of Biological Chemistry},
number = ,
volume = 281,
place = {United States},
year = {Sun Jan 01 00:00:00 EST 2006},
month = {Sun Jan 01 00:00:00 EST 2006}
}
  • No abstract prepared.
  • Carbon dioxide capture technologies have the potential to become an important climate change mitigation option through sequestration of gaseous CO 2, A new concept for CO 2 capture involves use of immobilized carbonic anhydrase (CA) that catalyzes the reversible hydration of CO 2 to HCO 3- and H +. Cost-efficient production of the enzyme and an inexpensive immobilization system are critical for development of economically feasible CA-based CO 2 capture processes. An artificial, bifunctional enzyme containing CA from Neisseria gonorrhoeae and a cellulose binding domain (CBD) from Clostridium thermocellum was constructed with a His 6 tag. The chimeric enzyme exhibitedmore » both CA activity and CBD binding affinity. This fusion enzyme is of particular interest due to its binding affinity for cellulose and retained CA activity, which could serve as the basis for improved technology to capture CO 2 from flue gasses.« less
  • Oxidation of methionine residues by reactive oxygen (ROS) in protein structures leads to the formation of methionine sulfoxide which can consequently lead to a plethora of impaired functionality. The generation of methionine sulfoxide yields ultimately a diastereomeric mixture of the S and R sulfoxides. So far two distinct enzyme families have been identified. MSRA reduces methionine S-sulfoxide, while MSRB reduces the R-diastereomer. It has been shown that these enzymes are involved in regulation of protein function and in elimination of ROS via reversible methionine formation besides protein repair. Importantly, both enzymes require coupling to the NADPH/thioredoxin reductase/thioredoxin electron donor system.more » In this report, we show for First time the expression and function of both sulfoxide reductases together with thioredoxin reductase in the cytosol as well as in the nucleus of epidermal melanocytes which are especially sensitive to ROS. Since this cell resides in the basal layer of the epidermis and its numbers and functions are reduced upon ageing and for instance also in depigmentation processes, we believe that this discovery adds an intricate repair mechanism to melanocyte homeostasis and survival.« less
  • The outer membrane proteins of Neisseria gonorrhoeae are specifically labeled by use of 1,3,4,6-tetrachloro-3..cap alpha..,6..cap alpha..-diphenyl glycoluril (Iodo-Gen) and /sup 125/I under the conditions described in this report. Use of this procedure with whole cells of N. gonorrhoeae produces a clear labeling pattern which can be visualized by electrophoretic separation of the proteins, followed by autoradiography. Electrophoretograms reveal some 70 polypeptide bands, while autoradiograms reveal only 5 or 6 labeled bands. The labeled polypeptide bands correspond to isolated outer membrane proteins, the most intensely labeled of which is the principal outer membrane protein. The method described in this report ismore » both specific and gentle, as well as rapid and convenient.« less