A specific stain for the detection of nonheme iron proteins in polyacrylamide gels.
Abstract: Nonheme iron proteins can be visualized as blue bands in native polyacrylamide gels using a staining method that is both simple and rapid. The reaction of potassium ferricyanide with protein-bound iron atoms to form royal blue complexes occurs almost instantaneously and is sensitive enough to detect 1 microgram of analytical-grade ferritin and 2 micrograms of purified ferredoxin from cyanobacteria. No special treatment of reagents or apparatus was necessary. On comparison, this stain was found to be more specific than the Ferene S stain, not detecting bovine serum albumin even when present as a hundredfold excess over ferritin. The method was found to be effective for isoelectric focusing gels as well.
Publication Date: 1992-12-01 PubMed ID: 1282787DOI: 10.1016/0003-2697(92)90018-3Google Scholar: Lookup
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- Comparative Study
- Journal Article
- Research Support
- Non-U.S. Gov't
Summary
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This research article presents a simple and quick method to visualize nonheme iron proteins in polyacrylamide gels. The staining technique uses potassium ferricyanide to reveal these proteins as blue bands on the gel, and has been found to be effective for isoelectric focusing gels as well.
Introduction
- The scientists explore a staining procedure that enables the detection of nonheme iron proteins in polyacrylamide gels.
- Nonheme iron proteins, unlike their heme counterparts, contain iron ions that are not bound to a heme group, yet, are critical for many biological functions. Detecting such proteins is therefore of utmost importance in biochemistry.
Procedure and Results
- The staining method used in this study employed potassium ferricyanide.
- This chemical reacts with the iron atoms bound to proteins to form royal blue complexes that are visible as blue bands in the gel.
- The researchers found the reaction to occur almost instantaneously and to be sensitive enough to detect minute quantities of certain iron-containing proteins, namely analytical-grade ferritin and purified ferredoxin from cyanobacteria.
- This procedure required no special treatment of reagents or apparatus.
Comparison with Other Staining Techniques and Applicabilities
- The study compared the effectiveness of this staining method with the commonly used Ferene S stain and found their method to be more specific.
- Unlike the Ferene S stain, this method failed to detect bovine serum albumin even when it was present in significant excess over ferritin, indicating its specificity for nonheme iron proteins.
- Additionally, this staining method was found to be effective for isoelectric focusing gels. Isoelectric focusing is a technique used to separate different proteins by their isoelectric point, and thus being able to employ this stain in the process would allow for greater analysis of nonheme iron proteins.
Cite This Article
APA
Leong LM, Tan BH, Ho KK.
(1992).
A specific stain for the detection of nonheme iron proteins in polyacrylamide gels.
Anal Biochem, 207(2), 317-320.
https://doi.org/10.1016/0003-2697(92)90018-3 Publication
Researcher Affiliations
- Department of Botany, Faculty of Science, National University of Singapore.
MeSH Terms
- Animals
- Blood Proteins / analysis
- Blood Proteins / isolation & purification
- Carrier Proteins / analysis
- Carrier Proteins / isolation & purification
- Cattle
- Chromogenic Compounds
- Electrophoresis, Polyacrylamide Gel / methods
- Ferredoxins / analysis
- Ferredoxins / isolation & purification
- Ferricyanides
- Ferritins / analysis
- Ferritins / isolation & purification
- Fetal Blood / chemistry
- Horses
- Humans
- Iron / analysis
- Iron-Binding Proteins
- Isoelectric Focusing / methods
- Microchemistry / methods
- Myoglobin / analysis
- Myoglobin / isolation & purification
- Rosaniline Dyes
- Serum Albumin, Bovine / analysis
- Serum Albumin, Bovine / isolation & purification
- Staining and Labeling
- Transferrin / analysis
- Transferrin / isolation & purification
- Transferrin-Binding Proteins
- Triazines
Citations
This article has been cited 17 times.- So M, Stiban J, Ciesielski GL, Hovde SL, Kaguni LS. Implications of Membrane Binding by the Fe-S Cluster-Containing N-Terminal Domain in the Drosophila Mitochondrial Replicative DNA Helicase.. Front Genet 2021;12:790521.
- Adameyko KI, Burakov AV, Finoshin AD, Mikhailov KV, Kravchuk OI, Kozlova OS, Gornostaev NG, Cherkasov AV, Erokhov PA, Indeykina MI, Bugrova AE, Kononikhin AS, Moiseenko AV, Sokolova OS, Bonchuk AN, Zhegalova IV, Georgiev AA, Mikhailov VS, Gogoleva NE, Gazizova GR, Shagimardanova EI, Gusev OA, Lyupina YV. Conservative and Atypical Ferritins of Sponges.. Int J Mol Sci 2021 Aug 11;22(16).
- Murgas CJ, Green SP, Forney AK, Korba RM, An SS, Kitten T, Lucas HR. Intracellular Metal Speciation in Streptococcus sanguinis Establishes SsaACB as Critical for Redox Maintenance.. ACS Infect Dis 2020 Jul 10;6(7):1906-1921.
- Fu H, Liu L, Dong Z, Guo S, Gao H. Dissociation between Iron and Heme Biosyntheses Is Largely Accountable for Respiration Defects of Shewanella oneidensis fur Mutants.. Appl Environ Microbiol 2018 Apr 15;84(8).
- Seim GL, Tako E, Ahn C, Glahn RP, Young SL. A Novel in Vivo Model for Assessing the Impact of Geophagic Earth on Iron Status.. Nutrients 2016 Jun 13;8(6).
- Hitchings MD, Townsend P, Pohl E, Facey PD, Jones DH, Dyson PJ, Del Sol R. A tale of tails: deciphering the contribution of terminal tails to the biochemical properties of two Dps proteins from Streptomyces coelicolor.. Cell Mol Life Sci 2014 Dec;71(24):4911-26.
- Williams SM, Chandran AV, Vijayabaskar MS, Roy S, Balaram H, Vishveshwara S, Vijayan M, Chatterji D. A histidine aspartate ionic lock gates the iron passage in miniferritins from Mycobacterium smegmatis.. J Biol Chem 2014 Apr 18;289(16):11042-11058.
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- Kawakami R, Sakuraba H, Ohshima T. Gene and primary structures of dye-linked L-proline dehydrogenase from the hyperthermophilic archaeon Thermococcus profundus show the presence of a novel heterotetrameric amino acid dehydrogenase complex.. Extremophiles 2004 Apr;8(2):99-108.
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