Metallomics Study of Lead-Protein Interactions in Albumen by
Electrochemical and Electrophoretic Methods

J 2012

Metallomics Study of Lead-Protein Interactions in Albumen by Electrochemical and Electrophoretic Methods

HYNEK, David, Ludmila KREJČOVÁ, Soňa KŘÍŽKOVÁ, Branislav RUTTKAY-NEDECKY, Jiri PIKULA et. al.

Basic information

Original name

Metallomics Study of Lead-Protein Interactions in Albumen by Electrochemical and Electrophoretic Methods

Authors

HYNEK, David (203 Czech Republic), Ludmila KREJČOVÁ (203 Czech Republic), Soňa KŘÍŽKOVÁ (203 Czech Republic), Branislav RUTTKAY-NEDECKY (203 Czech Republic), Jiri PIKULA (203 Czech Republic), Vojtěch ADAM (203 Czech Republic), Pavlina HAJKOVA (203 Czech Republic), Libuše TRNKOVÁ (203 Czech Republic, belonging to the institution), Jiri SOCHOR (203 Czech Republic), Miroslav POHANKA (203 Czech Republic), Jaromír HUBÁLEK (203 Czech Republic), Miroslava BEKLOVA (203 Czech Republic), Radimír VRBA (203 Czech Republic) and René KIZEK (203 Czech Republic, guarantor)

Edition

International Journal of Electrochemical Science, 2012, 1452-3981

Other information

Language

English

Type of outcome

Článek v odborném periodiku

Field of Study

10600 1.6 Biological sciences

Country of publisher

Serbia

Confidentiality degree

není předmětem státního či obchodního tajemství

Impact factor

Impact factor: 3.729 in 2011

RIV identification code

RIV/00216224:14310/12:00060305

Organization unit

Faculty of Science

UT WoS

000302730000004

Keywords in English

protein interaction; automated electrochemical detection; differential pulse voltammetry; heavy metal; capillary electrophoresis; metallothionein; spectrophotometry; proteomics

Abstract

V originále

Lead(II) ions represent still threat to living organisms due to high burden of these ions in environment. The aim of this study was to design and perform experiments studying the interaction of lead(II) ions and proteins contained in egg albumen. For this purpose, we used electrochemical and electrophoretic techniques. We focused on a very detailed study of the interaction of lead(II) ions with the egg albumen proteins. Firstly, egg albumen was diluted with acetate buffer in the following ratios buffer: albumen - 1:0.5; 1: 1; 1: 2 and 1: 4. Additions of different concentrations of lead(II) ions into the diluted egg albumen were 0; 1; 5; 10; 25; 50; 100; 150; 200; 250; 500; 750 and 1,000 nM. Mixtures were then placed on thermoblock at 37 degrees C for 15, 30, 45 and 60 min. Primarily, we used differential pulse anodic stripping voltammetry to analyse above prepared mixtures. It clearly follows from the obtained experimental data that there is a very strong interactions of lead(II) ions and biomolecules contained in egg albumen. Rapid increase in the concentration of lead(II) ions with a linear trend was detected in the mostly diluted egg albumen (lower concentration of total protein). At higher concentrations and longer time of interactions, metal ions bind tightly or intercalate into biomolecules presented in egg albumen. Besides the effect of lead(II) ions and albumen concentrations, we aimed at the effect of various times of incubation. The obtained results were linearly plotted and the slopes of these straight lines were compared. All slopes steep decreased with the increasing time of interaction and content of egg albumen proteins (15 min interaction, the ratio of 1: 4 and the slope 0.06; the ratio of 1: 2 and the slope 0.038; the ratio of 1: 1 and the slope 0.009; the ratio of 1: 0.5 and the slope 0.0085). Moreover, it is evident a clear time-dependence of binding lead(II) ions to biomolecules. At the ratio of 1: 0.5 there was observed the bond of app. 0.06 ng Pb per min. into biomolecules of egg albumen, at the ratio of 1: 1 there was observed the bond of app. 0.08 ng Pb per min. into biomolecules of egg albumen, at the ratio of 1: 2 there was observed the bond of 0.3 ng Pb per min. into biomolecules of egg albumen and at the ratio of 1: 4 there was observed the bond of 0.55 ng Pb per min. to biomolecules of egg albumen. Probably, in the case of lower concentrations of albumen proteins, there is a change in the structure of these proteins. The samples obtained by incubation of various concentrations of lead(II) ions with egg albumen protein were also analysed by chip capillary electrophoresis under non-reducing conditions to study the influence of lead(II) ions on the protein profiles of egg albumen. We determined changes in the content of the selected proteins as lysozyme, flavoprotein, ovalbumin, ovomucoid, avidin and ovotransferrin and some unidentified as marked according to their molecular masses 55, 95, 110, 117, 123, 132 and 170 kDa. Based on determined protein profiles (as slopes of signal changes depending on the applied lead(II) concentration) at various dilutions it is clearly evident that the most significant changes are detected at ovotransferrin, avidin, lysozyme and ovomucoid. In addition, we employed optimized methods for studying of lead(II) induced complexes in died embryo of vulture. Contents of lead(II) ions, reduced and oxidised glutathione, metallothionein and zinc(II) ions were determined in albumen, yolk, liver, kidney, brain and bone obtained from the embryo. The highest lead contents were observed in liver and kidney app. 5 mu g/g tissues. The highest contents of GSH and GSSH were found in brain tissue and in albumen, which can be associated with the needs to protect neural system against reactive oxygen radicals.

Links

MSM0021622412, plan (intention)

Name: Interakce mezi chemickými látkami, prostředím a biologickými systémy a jejich důsledky na globální, regionální a lokální úrovni (INCHEMBIOL) (Acronym: INCHEMBIOL)

Investor: Ministry of Education, Youth and Sports of the CR, Interactions among the chemicals, environment and biological systems and their consequences on the global, regional and local scales (INCHEMBIOL)

Citovat

HYNEK, David, Ludmila KREJČOVÁ, Soňa KŘÍŽKOVÁ, Branislav RUTTKAY-NEDECKY, Jiri PIKULA, Vojtěch ADAM, Pavlina HAJKOVA, Libuše TRNKOVÁ, Jiri SOCHOR, Miroslav POHANKA, Jaromír HUBÁLEK, Miroslava BEKLOVA, Radimír VRBA and René KIZEK. Metallomics Study of Lead-Protein Interactions in Albumen by Electrochemical and Electrophoretic Methods. International Journal of Electrochemical Science. 2012, vol. 7, No 2, p. 943-964. ISSN 1452-3981.

@article{986076,
   author = {Hynek, David and Krejčová, Ludmila and Křížková, Soňa and RuttkayandNedecky, Branislav and Pikula, Jiri and Adam, Vojtěch and Hajkova, Pavlina and Trnková, Libuše and Sochor, Jiri and Pohanka, Miroslav and Hubálek, Jaromír and Beklova, Miroslava and Vrba, Radimír and Kizek, René},
   article_number = {2},
   keywords = {protein interaction; automated electrochemical detection; differential pulse voltammetry; heavy metal; capillary electrophoresis; metallothionein; spectrophotometry; proteomics},
   language = {eng},
   issn = {1452-3981},
   journal = {International Journal of Electrochemical Science},
   title = {Metallomics Study of Lead-Protein Interactions in Albumen by Electrochemical and Electrophoretic Methods},
   volume = {7},
   year = {2012}
}

TY  - JOUR
ID  - 986076
AU  - Hynek, David - Krejčová, Ludmila - Křížková, Soňa - Ruttkay-Nedecky, Branislav - Pikula, Jiri - Adam, Vojtěch - Hajkova, Pavlina - Trnková, Libuše - Sochor, Jiri - Pohanka, Miroslav - Hubálek, Jaromír - Beklova, Miroslava - Vrba, Radimír - Kizek, René
PY  - 2012
TI  - Metallomics Study of Lead-Protein Interactions in Albumen by Electrochemical and Electrophoretic Methods
JF  - International Journal of Electrochemical Science
VL  - 7
IS  - 2
SP  - 943-964
EP  - 943-964
SN  - 14523981
KW  - protein interaction
KW  - automated electrochemical detection
KW  - differential pulse voltammetry
KW  - heavy metal
KW  - capillary electrophoresis
KW  - metallothionein
KW  - spectrophotometry
KW  - proteomics
N2  - Lead(II) ions represent still threat to living organisms due to high burden of these ions in environment. The aim of this study was to design and perform experiments studying the interaction of lead(II) ions and proteins contained in egg albumen. For this purpose, we used electrochemical and electrophoretic techniques. We focused on a very detailed study of the interaction of lead(II) ions with the egg albumen proteins. Firstly, egg albumen was diluted with acetate buffer in the following ratios buffer: albumen - 1:0.5; 1: 1; 1: 2 and 1: 4. Additions of different concentrations of lead(II) ions into the diluted egg albumen were 0; 1; 5; 10; 25; 50; 100; 150; 200; 250; 500; 750 and 1,000 nM. Mixtures were then placed on thermoblock at 37 degrees C for 15, 30, 45 and 60 min. Primarily, we used differential pulse anodic stripping voltammetry to analyse above prepared mixtures. It clearly follows from the obtained experimental data that there is a very strong interactions of lead(II) ions and biomolecules contained in egg albumen. Rapid increase in the concentration of lead(II) ions with a linear trend was detected in the mostly diluted egg albumen (lower concentration of total protein). At higher concentrations and longer time of interactions, metal ions bind tightly or intercalate into biomolecules presented in egg albumen. Besides the effect of lead(II) ions and albumen concentrations, we aimed at the effect of various times of incubation. The obtained results were linearly plotted and the slopes of these straight lines were compared. All slopes steep decreased with the increasing time of interaction and content of egg albumen proteins (15 min interaction, the ratio of 1: 4 and the slope 0.06; the ratio of 1: 2 and the slope 0.038; the ratio of 1: 1 and the slope 0.009; the ratio of 1: 0.5 and the slope 0.0085). Moreover, it is evident a clear time-dependence of binding lead(II) ions to biomolecules. At the ratio of 1: 0.5 there was observed the bond of app. 0.06 ng Pb per min. into biomolecules of egg albumen, at the ratio of 1: 1 there was observed the bond of app. 0.08 ng Pb per min. into biomolecules of egg albumen, at the ratio of 1: 2 there was observed the bond of 0.3 ng Pb per min. into biomolecules of egg albumen and at the ratio of 1: 4 there was observed the bond of 0.55 ng Pb per min. to biomolecules of egg albumen. Probably, in the case of lower concentrations of albumen proteins, there is a change in the structure of these proteins. The samples obtained by incubation of various concentrations of lead(II) ions with egg albumen protein were also analysed by chip capillary electrophoresis under non-reducing conditions to study the influence of lead(II) ions on the protein profiles of egg albumen. We determined changes in the content of the selected proteins as lysozyme, flavoprotein, ovalbumin, ovomucoid, avidin and ovotransferrin and some unidentified as marked according to their molecular masses 55, 95, 110, 117, 123, 132 and 170 kDa. Based on determined protein profiles (as slopes of signal changes depending on the applied lead(II) concentration) at various dilutions it is clearly evident that the most significant changes are detected at ovotransferrin, avidin, lysozyme and ovomucoid. In addition, we employed optimized methods for studying of lead(II) induced complexes in died embryo of vulture. Contents of lead(II) ions, reduced and oxidised glutathione, metallothionein and zinc(II) ions were determined in albumen, yolk, liver, kidney, brain and bone obtained from the embryo. The highest lead contents were observed in liver and kidney app. 5 mu g/g tissues. The highest contents of GSH and GSSH were found in brain tissue and in albumen, which can be associated with the needs to protect neural system against reactive oxygen radicals.
ER  -

HYNEK, David, Ludmila KREJČOVÁ, Soňa KŘÍŽKOVÁ, Branislav RUTTKAY-NEDECKY, Jiri PIKULA, Vojtěch ADAM, Pavlina HAJKOVA, Libuše TRNKOVÁ, Jiri SOCHOR, Miroslav POHANKA, Jaromír HUBÁLEK, Miroslava BEKLOVA, Radimír VRBA and René KIZEK. Metallomics Study of Lead-Protein Interactions in Albumen by Electrochemical and Electrophoretic Methods. \textit{International Journal of Electrochemical Science}. 2012, vol.~7, No~2, p.~943-964. ISSN~1452-3981.

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