Prefractionation by digitonin extraction increases representation of the cytosolic and intracellular proteome of Leishmania infantum

被引：41

作者：

Foucher, Aude L. ^{[1
]}

Papadopoulou, Barbara ^{[1
]}

Ouellette, Marc ^{[1
]}

机构：

[1] CHU Laval, Ctr Rech Infectiol, Quebec City, PQ G1V 4G2, Canada

来源：

JOURNAL OF PROTEOME RESEARCH | 2006年 / 5卷 / 07期

关键词：

fractionation; proteome; localization; Leishmania infantum;

D O I：

10.1021/pr060081j

中图分类号：

Q5 [生物化学];

学科分类号：

071010 ; 081704 ;

摘要：

Proteome coverage is limited by the dynamic range of proteins present in a sample and often is confined to the analysis of abundant proteins. We have developed a protein prefractionation protocol, based on the differential solubilization of membranes using digitonin, that has allowed an increase in the resolution and depth of comparative proteomic studies. This prefractionation protocol can also be used to infer the subcellular localization of hypothetical proteins as tested experimentally using green fluorescent fusion proteins. The abundant tubulins and associated proteins of the cytoskeleton were removed from the sample using digitonin extraction, hence facilitating the visualization of lower abundance proteins. The digitonin prefractionation protocol was applied for a comparative proteomic analysis of the promastigote and amastigote life cycle stages of Leishmania infantum and has allowed the identification of novel proteins expressed in a stage-specific manner.

引用

页码：1741 / 1750

页数：10

共 38 条

[1] Expression profiling of the Leishmania life cycle:: cDNA arrays identify developmentally regulated genes present but not annotated in the genome [J].

Almeida, R ;

Gilmartin, BJ ;

McCann, SH ;

Norrish, A ;

Ivens, AC ;

Lawson, D ;

Levick, MP ;

Smith, DF ;

Dyall, SD ;

Vetrie, D ;

Freeman, TC ;

Coulson, RM ;

Sampaio, I ;

Schneider, H ;

Blackwell, JM .

MOLECULAR AND BIOCHEMICAL PARASITOLOGY, 2004, 136 (01) :87-100

[2]

[Anonymous], 1988, Antibodies: A Laboratory Manual

[3] The Trypanosoma cruzi proteome [J].

Atwood, JA ;

Weatherly, DB ;

Minning, TA ;

Bundy, B ;

Cavola, C ;

Opperdoes, FR ;

Orlando, R ;

Tarleton, RL .

SCIENCE, 2005, 309 (5733) :473-476

[4] Developmentally induced changes of the proteome in the protozoan parasite Leishmania donovani [J].

Bente, M ;

Harder, S ;

Wiesgigl, M ;

Heukeshoven, J ;

Gelhaus, C ;

Krause, E ;

Clos, J ;

Bruchhaus, I .

PROTEOMICS, 2003, 3 (09) :1811-1829

[5] MEASUREMENT OF ACETYL-COA CARBOXYLASE ACTIVITY IN ISOLATED HEPATOCYTES [J].

BIJLEVELD, C ;

GEELEN, MJH .

BIOCHIMICA ET BIOPHYSICA ACTA, 1987, 918 (03) :274-283

[6] Large-scale identification of tubulin-binding proteins provides insight on subcellular trafficking, metabolic channeling, and signaling in plant cells [J].

Chuong, SDX ;

Good, AG ;

Taylor, GJ ;

Freeman, MC ;

Moorhead, GBG ;

Muench, DG .

MOLECULAR & CELLULAR PROTEOMICS, 2004, 3 (10) :970-983

[7] ENZYMIC CHARACTERIZATION AND LIPID COMPOSITION OF RAT LIVER SUBCELLULAR MEMBRANES [J].

COLBEAU, A ;

NACHBAUR, J ;

VIGNAIS, PM .

BIOCHIMICA ET BIOPHYSICA ACTA, 1971, 249 (02) :462-+

[8] Differential protein expression analysis of Leishmania major reveals novel roles for methionine adenosyltransferase and S-adenosylmethionine in methotrexate resistance [J].

Drummelsmith, J ;

Girard, I ;

Trudel, N ;

Ouellette, M .

JOURNAL OF BIOLOGICAL CHEMISTRY, 2004, 279 (32) :33273-33280

[9] Proteome mapping of the protozoan parasite Leishmania and application to the study of drug targets and resistance mechanisms [J].

Drummelsmith, J ;

Brochu, V ;

Girard, I ;

Messier, N ;

Ouellette, M .

MOLECULAR & CELLULAR PROTEOMICS, 2003, 2 (03) :146-155

[10]

El Fakhry Y, 2002, PROTEOMICS, V2, P1007, DOI 10.1002/1615-9861(200208)2:8<1007::AID-PROT1007>3.0.CO

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