Biocytin
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| Surname | Biocytin | ||||||||||||
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| Molecular formula | C 16 H 28 N 4 O 4 S | ||||||||||||
| Brief description |
beige powder |
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| properties | |||||||||||||
| Molar mass | 372.48 g mol −1 | ||||||||||||
| Physical state |
firmly |
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| Melting point |
245 ° C |
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| As far as possible and customary, SI units are used. Unless otherwise noted, the data given apply to standard conditions . | |||||||||||||
Biocytin or N ε - (+) - biotinyl- L -lysine is a chemical compound that belongs to the carboxamides . The common name biocytin is used exclusively for the combination of the vitamin biotin [(+) - stereoisomer ] and the L - enantiomer of the amino acid lysine . The is carboxyl group of biotin with the ε- amino group of lysine condensed . As an intermediate product of the biotin metabolism, biocytin occurs in the blood serum and in the urine .
Since the late 1980s, biocytin has been used as a so-called neuronal tracer as a component of histological staining methods for nerve cells .
Occurrence and biological importance
Biocytin is the biotin- bearing complex in the following carboxylases , which perform basic tasks in carbohydrate, protein and fat metabolism:
- Pyruvate carboxylase
- Methyl crotonoyl-CoA carboxylase
- Propionyl-CoA carboxylase
- Acetyl-CoA carboxylase
When these carboxylases are broken down by proteolysis , biocytin is left over. The cleavage of the biocytin into free biotin and lysine is catalyzed by the enzyme biotinidase . Since animals cannot produce the vitamin biotin themselves, they are dependent on ingesting biotin with their food and recycling biotin already present in the body many times over. Therefore, biocytin is an important source of biotin for the organism.
The importance of biocytin for humans in this regard is made clear by the rare hereditary disease biotinidase deficiency . Due to a defect in the biotinidase, those affected cannot use biocytin sufficiently and develop symptoms of a biotin deficiency as a result, which can lead to death.
properties
Commercially available biocytin is a beige powder that melts at 245 ° C. Like biotin, biocytin is not covalently bound by avidin .
The enzyme biotinidase hydrolyzes the amide bond of the biocytin to form biotin and lysine.
use
Biotinidase test
Since biocytin is the natural substrate of the enzyme biotinidase , it can be used to measure biotinidase activity. The amount of the reaction products biotin or lysine formed per unit of time is a measure of the activity of the enzyme. For the determination of the biotinidase activity in human blood samples, in particular for the diagnosis of the metabolic disorder biotinidase deficiency, biocytin is only of limited suitability. The medication with biotin can the enzyme defect disguise.
Neural tracer
Loading with biocytin is one of many ways to make neurons, including their widely ramified branches, visible. It is possible to apply biocytin to living tissue, where it is distributed over the nerve cells within hours. Individual neurons can also be filled with biocytin via microinjection. After this step, the neurons are marked, but not yet stained because biocytin itself is not a dye. The color of the adhesive Biocytins place after washing and fusing procedure at the intersection preparation instead. This makes use of the fact that the protein avidin binds very strongly to biotin and biotin compounds such as biocytin. For example, a fluorescent dye can be biotinylated and then allowed to dock onto avidin. Since avidin has several binding sites for biotin structures, the avidin-fluorophore complex can in turn dock onto the biocytin that marks the nerve cell - the cell fluoresces. Another possibility is to allow an avidin- peroxidase complex to attach to the biocytin. However, this is not yet colored. One color variant consists in a reaction of the peroxidase with diaminobenzidine (DAB), hydrogen peroxide and divalent nickel and / or cobalt ions.
As the two examples show, when using biocytin to mark neurons, the actual staining method can be used very flexibly. It is thus possible to produce suitable preparations for light microscopy , fluorescence microscopy and electron microscopy .
safety instructions
According to a distributor , no data are currently available on the toxicity of biocytin, but the substance has not yet been fully tested. According to the CLP regulation this substance is not classified as dangerous. When working with the pure substance, generally accepted precautionary measures are recommended. Due to the self-classification of the manufacturer, biocytin is in water hazard class (WGK) 3.
Individual evidence
- ↑ a b c d e f sheet biocytin at Sigma-Aldrich , retrieved on May 22, 2017 ( PDF ).
- ↑ JM Berg, JL Tymoczko, L. Stryer: Biochemistry. 6th edition. Spectrum Academic Publishing House, Elsevier GmbH, Munich 2007; Pp. 515-516, 551, 697-698, 711-713, 746, ISBN 978-3-8274-1800-5 .
- ↑ ER Baumgartner, T. Suormala: Multiple carboxylase deficiency: inherited and acquired disorders of biotin metabolism. In: Int. J. Vitam. Nutr. Res. 67 (5); 1997: pp. 377-784; PMID 9350481 .
- ↑ M. Koivusalo, C. Elorriaga, Y. Kaziro, S. Ochoa: Bacterial biotinidase. In: J. Biol. Chem. 238; March 1963: pp. 1038-1042; PMID 14034272 .
- ↑ K. Kumasaka, M. Muratsugu, T. Fukui, M. Kimura, Y. Takagi, N. Hashizume: A new quantitative analytical method of serum biotinidase activity using biocytin as a substrate and its clinical significance in Japan. In: Clin. Chim. Acta 306 (1-2); April 2001: pp. 71-77; PMID 11282096 ; doi : 10.1016 / S0009-8981 (01) 00395-3
- ↑ C. Köbbert, R. Apps, I. Bechmann, JL Lanciego, J. Mey, S. Thanos: Current concepts in neuroanatomical tracing. In: Prog. Neurobiol. 62 (4); Nov 2000: pp. 327-351, PMID 10856608 ; doi : 10.1016 / S0301-0082 (00) 00019-8
- ^ AJ McDonald: Neuroanatomical labeling with biocytin: a review. In: Neuroreport 3 (10); Oct 1992: pp. 821-827; PMID 1384763 .