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Crystallization and preliminary X-ray diffraction analysis of eukaryotic α2-macroglobulin family members modified by methylamine, proteases and glycosidases

Published on Dec 1, 2014in Molecular Oral Microbiology2.925
· DOI :10.1111/omi.12069
Theodoros Goulas8
Estimated H-index: 8
,
I. Garcia-Ferrer6
Estimated H-index: 6
+ 2 AuthorsF.X. Gomis-Rüth1
Estimated H-index: 1
Abstract
European, Spanish, and Catalan Agencies. Grant Number: FP7-HEALTH-2010-261460; GumsJ RAPID. Grant Number: FP7-HEALTH-2012-306029-2; TRIGGER. Grant Numbers: BFU2012-32862, CSD2006-00015; La Marato de TV3. Grant Numbers: 2009-100732, 2009SGR1036; ESRF
  • References (40)
  • Citations (5)
References40
Newest
Last. Ina AttreeH-Index: 29
view all 8 authors...
ABSTRACT Human pathogens frequently use protein mimicry to manipulate host cells in order to promote their survival. Here we show that the opportunistic pathogen Pseudomonas aeruginosa synthesizes a structural homolog of the human α2-macroglobulin, a large-spectrum protease inhibitor and important player of innate immunity. Small-angle X-ray scattering analysis demonstrated that the fold of P. aeruginosa MagD (PA4489) is similar to that of the human macroglobulin and undergoes a conformational m...
15 CitationsSource
#1Ahmed Abdur Rehman (AMU: Aligarh Muslim University)H-Index: 6
#2Haseeb Ahsan (Jamia Millia Islamia)H-Index: 22
Last. Fahim Halim Khan (AMU: Aligarh Muslim University)H-Index: 9
view all 3 authors...
Alpha macroglobulins are large glycoproteins which are present in the body fluids of both invertebrates and vertebrates. Alpha-2-macroglobulin (α2M), a key member of alpha macroglobulin superfamily, is a high-molecular weight homotetrameric glycoprotein. α2M has many diversified and complex functions, but it is primarily known by its ability to inhibit a broad spectrum of proteases without the direct blockage of the protease active site. α2M is also known to be involved in the regulation, transp...
108 CitationsSource
#1Cindy Meyer (UHH: University of Hamburg)H-Index: 13
#2Winfried Hinrichs (University of Greifswald)H-Index: 30
Last. Ulrich Hahn (UHH: University of Hamburg)H-Index: 30
view all 3 authors...
15 CitationsSource
#1A. Marrero (CSIC: Spanish National Research Council)H-Index: 8
#2Stéphane Duquerroy (University of Paris-Sud)H-Index: 21
Last. F. Xavier Gomis-Rüth (CSIC: Spanish National Research Council)H-Index: 31
view all 9 authors...
61 CitationsSource
#1Syed R. Haider (Lboro: Loughborough University)H-Index: 4
#2Barry L. Sharp (Lboro: Loughborough University)H-Index: 21
Last. Helen J. Reid (Lboro: Loughborough University)H-Index: 14
view all 3 authors...
This paper compares different buffer systems for the electrophoretic separation of the five most abundant serum proteins on native-PAGE gel and cellulose membranes. A modified Tris-tricine system was shown to be superior for the separation of these serum proteins in a 7% m/v native-PAGE gel as compared with the traditionally used Tris-glycine and Tris-tricine methods. This modified Tris-tricine buffer system was also employed for the separation of serum proteins using a cellulose acetate membran...
10 CitationsSource
#1Johann Schaller (University of Bern)H-Index: 26
#2Simon S. Gerber (University of Bern)H-Index: 2
The plasmin–antiplasmin system plays a key role in blood coagulation and fibrinolysis. Plasmin and α2-antiplasmin are primarily responsible for a controlled and regulated dissolution of the fibrin polymers into soluble fragments. However, besides plasmin(ogen) and α2-antiplasmin the system contains a series of specific activators and inhibitors. The main physiological activators of plasminogen are tissue-type plasminogen activator, which is mainly involved in the dissolution of the fibrin polyme...
88 CitationsSource
#1Theodoros GoulasH-Index: 8
#2Joan L. ArolasH-Index: 17
Last. F. Xavier Gomis-Rüth (CSIC: Spanish National Research Council)H-Index: 31
view all 3 authors...
Enterotoxigenic Bacteroides fragilis is the most frequent disease-causing anaerobe in the intestinal tract of humans and livestock and its specific virulence factor is fragilysin, also known as B. fragilis toxin. This is a 21-kDa zinc-dependent metallopeptidase existing in three closely related isoforms that hydrolyze E-cadherin and contribute to secretory diarrhea, and possibly to inflammatory bowel disease and colorectal cancer. Here we studied the function and zymogenic structure of fragilysi...
39 CitationsSource
#1Tomasz Kantyka (Jagiellonian University)H-Index: 20
#2Neil D. Rawlings (Wellcome Trust Sanger Institute)H-Index: 43
Last. Jan Potempa (Jagiellonian University)H-Index: 63
view all 3 authors...
In metazoan organisms protein inhibitors of peptidases are important factors essential for regulation of proteolytic activity. In vertebrates genes encoding peptidase inhibitors constitute up to 1% of genes reflecting a need for tight and specific control of proteolysis especially in extracellular body fluids. In stark contrast unicellular organisms, both prokaryotic and eukaryotic consistently contain only few, if any, genes coding for putative peptidase inhibitors. This may seem perplexing in ...
31 CitationsSource
#1Mauricio M. PaivaH-Index: 6
#2Maria de Nazaré C. Soeiro (FIOCRUZ: Oswaldo Cruz Foundation)H-Index: 26
Last. T. C. Araujo-Jorge (FIOCRUZ: Oswaldo Cruz Foundation)H-Index: 11
view all 6 authors...
Abstract Human alpha2-macroglobulin (α2M) is a 720 kDa glycoprotein that presents two ultrastructural conformations: slow (S-α2M) and fast (F-α2M). α2M acts mainly as a proteinase scavenger, but an immunomodulatory role was also proposed. This work studies the effect of desialylation and deglycosylation on the structure patterns of α2M by ultrastructural analysis of lectin-induced aggregates, which represents a new approach that had never been previously used. Transmission electron microscopy (T...
3 CitationsSource
α-Macroglobulins (αMs) are large glycoproteins that have been identified in a wide range of vertebrate and invertebrate species and are mostly thiol ester containing proteinase inhibitors. A recent analysis of bacterial genomes (Budd, A., Blandin, S., Levashina, E. A., and Gibson, T. J. (2004) Genome Biol. 5 , R38.) identified many α-macroglobulin-like sequences that appear to have been acquired by Gram-negative bacteria from their metazoan hosts. We report the first expression and characterizat...
16 CitationsSource
Cited By5
Newest
#1L. Del Amo-MaestroH-Index: 1
Last. F.X. Gomis-RuthH-Index: 1
view all 4 authors...
Transforming growth factor β is a disulfide-linked dimeric cytokine that occurs in three highly related isoforms (TGFβ1–TGFβ3) engaged in signaling functions through binding of cognate TGFβ receptors. To regulate this pathway, the cytokines are biosynthesized as inactive pro-TGFβs with an N-terminal latency-associated protein preceding the mature moieties. Due to their pleiotropic implications in physiology and pathology, TGFβs are privileged objects of in vitro studies. However, such studies ha...
1 CitationsSource
#1Laura Marino-Puertas (CSIC: Spanish National Research Council)H-Index: 1
#2Laura del Amo-Maestro (CSIC: Spanish National Research Council)
Last. Theodoros Goulas (CSIC: Spanish National Research Council)H-Index: 8
view all 5 authors...
α2-Macroglobulins (α2Ms) regulate peptidases, hormones and cytokines. Mediated by peptidase cleavage, they transit between native, intact forms and activated, induced forms. α2Ms have been studied over decades using authentic material from primary sources, which was limited by sample heterogeneity and contaminants. Here, we developed high-yield expression systems based on transient transfection in Drosophila Schneider 2 and human Expi293F cells, which produced pure human α2M (hα2M) at ~1.0 and ~...
Source
#1Theodoros GoulasH-Index: 8
#2I. Garcia-FerrerH-Index: 6
Last. F. Xavier Gomis-RüthH-Index: 31
view all 6 authors...
This study was supported in part by grants from European, Spanish, and Catalan agencies (grant references FP7-HEALTH-2012-306029-2 ‘TRIGGER’, BFU2015-64487R, MDM-2014-0435, BIO2013-49320-EXP, and 2014SGR9). The Structural Biology Unit (www.sbu.csic.es) of IBMB has been named a ‘Maria de Maeztu’ Unit of Excellence by the Spanish Ministry of Economy and Competitiveness. T.G. and I.G.-F. acknowledge a ‘Juan-dela-Cierva’ contract from the Spanish Ministry of Economy and Competitiveness (JCI-2012-135...
7 CitationsSource
#1I. Garcia-Ferrer (CSIC: Spanish National Research Council)H-Index: 6
#2A. Marrero (CSIC: Spanish National Research Council)H-Index: 8
Last. Theodoros Goulas (CSIC: Spanish National Research Council)H-Index: 8
view all 4 authors...
α2-macroglobulins are broad-spectrum endopeptidase inhibitors, which have to date been characterised from metazoans (vertebrates and invertebrates) and Gram-negative bacteria. Their structural and biochemical properties reveal two related modes of action: the “Venus flytrap” and the “snap-trap” mechanisms. In both cases, peptidases trigger a massive conformational rearrangement of α2-macroglobulin after cutting in a highly flexible bait region, which results in their entrapment. In some homologs...
5 CitationsSource
#1Jan Potempa (University of Louisville)H-Index: 63
#2Richard J. Lamont (University of Louisville)H-Index: 59
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