Isolation and functional characterization of Sporothrix schenckii ROT2, the encoding gene for the endoplasmic reticulum glucosidase II

2012 ◽  
Vol 116 (8) ◽  
pp. 910-918 ◽  
Author(s):  
Claudia I. Robledo-Ortiz ◽  
Arturo Flores-Carreón ◽  
Arturo Hernández-Cervantes ◽  
Aurelio Álvarez-Vargas ◽  
Keunsook K. Lee ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Functional Characterization ◽  
Sporothrix Schenckii ◽  
Glucosidase Ii ◽  
Encoding Gene

scholarly journals N-Acetylglucosamine-1-Phosphate Transferase, WecA, as a Validated Drug Target in Mycobacterium tuberculosis

2017 ◽  
Vol 61 (11) ◽  
Author(s):  
Stanislav Huszár ◽  
Vinayak Singh ◽  
Alica Polčicová ◽  
Peter Baráth ◽  
María Belén Barrio ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Drug Target ◽  
Functional Characterization ◽  
Drug Candidate ◽  
Content Type ◽  
Chemical Libraries ◽  
Whole Cells ◽  
Encoding Gene

ABSTRACT The mycobacterial phosphoglycosyltransferase WecA, which initiates arabinogalactan biosynthesis in Mycobacterium tuberculosis, has been proposed as a target of the caprazamycin derivative CPZEN-45, a preclinical drug candidate for the treatment of tuberculosis. In this report, we describe the functional characterization of mycobacterial WecA and confirm the essentiality of its encoding gene in M. tuberculosis by demonstrating that the transcriptional silencing of wecA is bactericidal in vitro and in macrophages. Silencing wecA also conferred hypersensitivity of M. tuberculosis to the drug tunicamycin, confirming its target selectivity for WecA in whole cells. Simple radiometric assays performed with mycobacterial membranes and commercially available substrates allowed chemical validation of other putative WecA inhibitors and resolved their selectivity toward WecA versus another attractive cell wall target, translocase I, which catalyzes the first membrane step in the biosynthesis of peptidoglycan. These assays and the mutant strain described herein will be useful for identifying potential antitubercular leads by screening chemical libraries for novel WecA inhibitors.

Identification and functional characterization of a novel human protein highly related to the yeast dynamin-like GTPase Vps1p

1998 ◽  
Vol 111 (10) ◽  
pp. 1341-1349 ◽  
Author(s):  
M. Imoto ◽  
I. Tachibana ◽  
R. Urrutia
Keyword(s):  
Endoplasmic Reticulum ◽  
Cho Cells ◽  
Mammalian Cells ◽  
Functional Characterization ◽  
Luciferase Reporter ◽  
Pleckstrin Homology ◽  
Rich Region ◽  
Gtpase Domain ◽  
Proline Rich Region

Dynamin proteins containing a GTPase domain, a pleckstrin homology motif and a proline-rich tail participate in receptor-mediated endocytosis in organisms ranging from insects to vertebrates. In addition, dynamin-related GTPases, such as the yeast Golgi protein Vps1p, which lack both the pleckstrin homology motif and the proline-rich region, participate in vesicular transport within the secretory pathway in lower eukaryotes. However, no data is available on the existence of Vps1p-like proteins in mammalian cells. In this study, we report the identification and characterization of a novel gene encoding a human dynamin-related protein, DRP1, displaying high similarity to the Golgi dynamin-like protein Vps1p from yeast and to a Caenorhabditis elegans protein deposited in the databank. These proteins are highly conserved in their N-terminal tripartite GTPase domain but lack the pleckstrin homology motif and proline-rich region. Northern blot analysis reveals that the DRP1 mRNA is detected at high levels in human muscle, heart, kidney and brain. Immunolocalization studies in Chinese hamster ovary (CHO) cells using an epitope-tagged form of DRP1 and confocal microscopy show that this protein is concentrated in a perinuclear region that labels with the endoplasmic reticulum marker DiOC6(3) and the Golgi marker C5-DMB-Cer. In addition, the localization of DRP1 is highly similar to the localization of the endoplasmic reticulum and cis-Golgi GTPase Rab1A, but not to the staining for the trans-Golgi GTPase Rab6. Furthermore, overexpression of a cDNA encoding a GTP binding site mutant of DRP1 (DRP1(K38E)) in CHO cells decreases the amount of a secreted luciferase reporter protein, whereas the overexpression of wild-type DRP1 increases the secretion of this marker. Together, these results constitute the first structural and functional characterization of a mammalian protein similar to the yeast dynamin-related GTPase Vps1p and indicate that the participation of these proteins in secretion has been conserved throughout evolution.

scholarly journals Functional characterization of the oxaloacetase encoding gene and elimination of oxalate formation in the β-lactam producer Penicillium chrysogenum

2011 ◽  
Vol 48 (8) ◽  
pp. 831-839 ◽  
Author(s):  
A.K. Gombert ◽  
T. Veiga ◽  
M. Puig-Martinez ◽  
F. Lamboo ◽  
J.G. Nijland ◽  
...  
Keyword(s):  
Penicillium Chrysogenum ◽  
Functional Characterization ◽  
Encoding Gene

scholarly journals Functional Characterization of ERp18, a New Endoplasmic Reticulum-located Thioredoxin Superfamily Member

2003 ◽  
Vol 278 (31) ◽  
pp. 28912-28920 ◽  
Author(s):  
Heli I. Alanen ◽  
Richard A. Williamson ◽  
Mark J. Howard ◽  
Anna-Kaisa Lappi ◽  
Heli P. Jäntti ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Functional Characterization

Isolation of Sporothrix schenckii MNT1 and the biochemical and functional characterization of the encoded α1,2-mannosyltransferase activity

Microbiology ◽  
2012 ◽  
Vol 158 (9) ◽  
pp. 2419-2427 ◽  
Author(s):  
Arturo Hernández-Cervantes ◽  
Héctor M. Mora-Montes ◽  
Aurelio Álvarez-Vargas ◽  
Diana F. Díaz Jiménez ◽  
Claudia I. Robledo-Ortiz ◽  
...  
Keyword(s):  
Functional Characterization ◽  
Sporothrix Schenckii

Functional characterization of orchardgrass endoplasmic reticulum-resident Hsp90 (DgHsp90) as a chaperone and an ATPase

2009 ◽  
Vol 47 (10) ◽  
pp. 859-866 ◽  
Author(s):  
Joon-Yung Cha ◽  
Min Hee Jung ◽  
Netty Ermawati ◽  
Mukhamad Su'udi ◽  
Gyu-Jin Rho ◽  
...  
Keyword(s):  
Endoplasmic Reticulum ◽  
Functional Characterization

scholarly journals Structural and functional characterization of Sec66p, a new subunit of the polypeptide translocation apparatus in the yeast endoplasmic reticulum.

1993 ◽  
Vol 4 (9) ◽  
pp. 931-939 ◽  
Author(s):  
D Feldheim ◽  
K Yoshimura ◽  
A Admon ◽  
R Schekman
Keyword(s):  
Endoplasmic Reticulum ◽  
Signal Sequence ◽  
Functional Characterization ◽  
Yeast Cells ◽  
Temperature Sensitive ◽  
Endoplasmic Reticulum Membrane ◽  
Restrictive Temperature ◽  
Permissive Temperature

SEC66 encodes the 31.5-kDa glycoprotein of the Sec63p complex, an integral endoplasmic reticulum membrane protein complex required for translocation of presecretory proteins in Saccharomyces cerevisiae. DNA sequence analysis of SEC66 predicts a 23-kDa protein with no obvious NH2-terminal signal sequence but with one domain of sufficient length and hydrophobicity to span a lipid bilayer. Antibodies directed against a recombinant form of Sec66p were used to confirm the membrane location of Sec66p and that Sec66p is a glycoprotein of 31.5 kDa. A null mutation in SEC66 renders yeast cells temperature sensitive for growth. sec66 cells accumulate some secretory precursors at a permissive temperature and a variety of precursors at the restrictive temperature. sec66 cells show defects in Sec63p complex formation. Because sec66 cells affect the translocation of some, but not all secretory precursor polypeptides, the role of Sec66p may be to interact with the signal peptide of presecretory proteins.

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