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Title: Molecular genetic and biochemical characterization of a putative family of zinc metalloproteins in Caenorhabditis elegans

Abstract

Four highly similar genes ( W08E12.2, W08E12.3, W08E12.4 and W08E12.5) which are consecutively aligned on chromosome IV of the C. elegans genome are predicted to code for small (120–141aa) yet cysteine rich (18–19 cysteines) proteins. Cloning and sequencing of the genomic regions of the isoforms confirmed the presence and order of all genes. The generation of transgenic worms strains with an integrated single copy or extrachromosomal multi-copy P W08E12.3;W08E12.4::GFP uncovered that W08E12.3 and W08E12.4 are constitutively expressed in the pharynx and significantly induced in worms exposed to 100 μM Zn. Knockdown by RNAi did not have a marked consequence on reproductive performance nor was a Zn-dependent effect on nematode growth observed. However, RNAi of these genes led to an accumulation of Zn in the intestinal cells. W08E12.3 was recombinantly expressed in E. coli and the purified protein was shown to be able to bind up to 6.5 Zn molecules at neutral pH. Zn-binding was acid-labile and the apo protein was observed at pH < 4.3. In conclusion, this characterization suggests W08E12.2, W08E12.3, W08E12.4 and W08E12.5 belong to a family of putative Metalloproteins which, akin to metallothioneins, may play an important role in Zn-sensing, homeostasis and/or detoxification.

Authors:
 [1];  [2];  [1];  [1];  [3]; ORCiD logo [2]; ORCiD logo [1]
  1. King's College London, London (United Kingdom)
  2. Univ. of Warwick, Coventry (United Kingdom)
  3. SLAC National Accelerator Lab., Menlo Park, CA (United States)
Publication Date:
Research Org.:
SLAC National Accelerator Lab., Menlo Park, CA (United States)
Sponsoring Org.:
USDOE Office of Science (SC), Basic Energy Sciences (BES) (SC-22)
OSTI Identifier:
1482410
Alternate Identifier(s):
OSTI ID: 1490674
Grant/Contract Number:  
BB/E025064; BB/E025099; AC02-76SF00515
Resource Type:
Published Article
Journal Name:
Metallomics
Additional Journal Information:
Journal Volume: 10; Journal Issue: 12; Journal ID: ISSN 1756-5901
Publisher:
Royal Society of Chemistry
Country of Publication:
United States
Language:
English
Subject:
59 BASIC BIOLOGICAL SCIENCES

Citation Formats

Chaudhuri, Poulami, Imam, Hasan Tanvir, Essig, Yona, Krasauskas, Jovaras, Webb, Samuel M., Blindauer, Claudia A., and Stürzenbaum, Stephen R. Molecular genetic and biochemical characterization of a putative family of zinc metalloproteins in Caenorhabditis elegans. United States: N. p., 2018. Web. doi:10.1039/c8mt00169c.
Chaudhuri, Poulami, Imam, Hasan Tanvir, Essig, Yona, Krasauskas, Jovaras, Webb, Samuel M., Blindauer, Claudia A., & Stürzenbaum, Stephen R. Molecular genetic and biochemical characterization of a putative family of zinc metalloproteins in Caenorhabditis elegans. United States. doi:10.1039/c8mt00169c.
Chaudhuri, Poulami, Imam, Hasan Tanvir, Essig, Yona, Krasauskas, Jovaras, Webb, Samuel M., Blindauer, Claudia A., and Stürzenbaum, Stephen R. Fri . "Molecular genetic and biochemical characterization of a putative family of zinc metalloproteins in Caenorhabditis elegans". United States. doi:10.1039/c8mt00169c.
@article{osti_1482410,
title = {Molecular genetic and biochemical characterization of a putative family of zinc metalloproteins in Caenorhabditis elegans},
author = {Chaudhuri, Poulami and Imam, Hasan Tanvir and Essig, Yona and Krasauskas, Jovaras and Webb, Samuel M. and Blindauer, Claudia A. and Stürzenbaum, Stephen R.},
abstractNote = {Four highly similar genes (W08E12.2, W08E12.3, W08E12.4 and W08E12.5) which are consecutively aligned on chromosome IV of the C. elegans genome are predicted to code for small (120–141aa) yet cysteine rich (18–19 cysteines) proteins. Cloning and sequencing of the genomic regions of the isoforms confirmed the presence and order of all genes. The generation of transgenic worms strains with an integrated single copy or extrachromosomal multi-copy PW08E12.3;W08E12.4::GFP uncovered that W08E12.3 and W08E12.4 are constitutively expressed in the pharynx and significantly induced in worms exposed to 100 μM Zn. Knockdown by RNAi did not have a marked consequence on reproductive performance nor was a Zn-dependent effect on nematode growth observed. However, RNAi of these genes led to an accumulation of Zn in the intestinal cells. W08E12.3 was recombinantly expressed in E. coli and the purified protein was shown to be able to bind up to 6.5 Zn molecules at neutral pH. Zn-binding was acid-labile and the apo protein was observed at pH < 4.3. In conclusion, this characterization suggests W08E12.2, W08E12.3, W08E12.4 and W08E12.5 belong to a family of putative Metalloproteins which, akin to metallothioneins, may play an important role in Zn-sensing, homeostasis and/or detoxification.},
doi = {10.1039/c8mt00169c},
journal = {Metallomics},
number = 12,
volume = 10,
place = {United States},
year = {2018},
month = {11}
}

Journal Article:
Free Publicly Available Full Text
Publisher's Version of Record
DOI: 10.1039/c8mt00169c

Figures / Tables:

Fig. 1 Fig. 1: (A) Schematic diagram of a section of cosmid W08E12 which is part of chromosome IV depicting the location of W08E12.2, W08E12.3, W08E12.4 and W08E12.5. Note the coding sequences of W08E12.4 and W08E12.5 (red boxes), and the first 500 bp of the W08E12.3 and W08E12.4 promoters (red dashed lines)more » are identical. (B) ClustalW alignment of amino acid sequences of W08E12.2, W08E12.3, W08E12.4 and W08E12.5. Identical residues are shown in black and cysteines are highlighted in red. Note: the high level of sequence identity between W08E12.3, W08E12.4 and W08E12.5, with W08E12.2 being shorter and lacking the N terminal cysteine.« less

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    Figures/Tables have been extracted from DOE-funded journal article accepted manuscripts.