المرجع الالكتروني للمعلوماتية
المرجع الألكتروني للمعلوماتية

علم الاحياء
عدد المواضيع في هذا القسم 10456 موضوعاً
النبات
الحيوان
الأحياء المجهرية
علم الأمراض
التقانة الإحيائية
التقنية الحياتية النانوية
علم الأجنة
الأحياء الجزيئي
علم وظائف الأعضاء
المضادات الحيوية

Untitled Document
أبحث عن شيء أخر المرجع الالكتروني للمعلوماتية
من هم المحسنين؟
2024-11-23
ما هي المغفرة؟
2024-11-23
{ليس لك من الامر شيء}
2024-11-23
سبب غزوة أحد
2024-11-23
خير أئمة
2024-11-23
يجوز ان يشترك في الاضحية اكثر من واحد
2024-11-23

صحيفة الاهرام
4-3-2022
خصلتان ذميمتان لليهود
2024-08-23
Collagen Band-Ligament
8-1-2017
أمراض الجرب البكتيري
31-1-2023
رسم مسار الأشعة بالنسبة للعدسات الرقيقة ؛ معادلة العدسة الرقيقة
11-1-2016
خدمة اللوبيا بعد الزراعة
17-5-2018

Avidin-Biotin System  
  
3178   12:59 صباحاً   date: 9-12-2015
Author : M. Wilchek and E. A. Bayer, eds.
Book or Source : Avidin-Biotin Technology Methods in Enzymology
Page and Part :


Read More
Date: 1-12-2015 2332
Date: 7-5-2021 1658
Date: 18-5-2016 2033

 Avidin-Biotin System

 The avidin-biotin system has become one of the major mainstays in biochemical analysis and has far-reaching application in biotechnology, industry, and clinical medicine (1). The general idea of the avidin-biotin system is that biotin, a low molecular weight vitamin, can be chemically coupled to other low or high molecular weight molecules (e.g., proteins, hormones, DNA molecules, etc.). The biotin moiety is still recognized by avidin or streptavidin, either as the native protein or in derivatized form containing any one of a number of reporter groups or probes. The principle of this system is illustrated in Fig. 1

Figure 1. Overview of the avidin-biotin system and the two major strategies for the various applications. In both approach desired experimental system is combined with a biotinylated binder molecule. Approach A involves direct interaction with probe. In approach B, avidin is a sandwich between the biotinylated binder and the biotinylated probe. Various targets, b the applications are listed.

The avidin-biotin system has become a “universal” tool in most of the fields of the biological sciences, because of studies that commenced in the mid 1970s and the constant development that has continued until today (2-6). The system has been applied for a wide variety of purposes (Fig. 1) and has recently been adapted for clinical use for the localization, imaging, and therapy of cancer (7). The binder and the target can be any of the components listed in Figure 1. What is required for this system is the capacity to biotinylate a binding entity so that the specificity and activity of the binding function is retained. For different approaches to biotinylation and the reagents used for binding to different functional groups. As can be seen, most of the functional groups on biological molecules can be modified with biotin. Because of its charge neutrality and lack of glycosylation, streptavidin is generally preferred over egg-white avidin in many applications, although new derivatives of avidin (e.g., Neutravidin) may prove advantageous and less expensive. The final component added to the system is the probe. The various probes and their potential uses are shown in Fig. 1. The probes are prepared in two ways. They are chemically conjugated directly to avidin or streptavidin (Fig. 1, Approach A) and are fluorescent, radioactive, or other types of macromolecules (proteins, polysaccharides, etc.). A second approach is to biotinylate the probes and to interact them with streptavidin under subsaturating ratios, thus leaving extra binding sites vacant (Fig. 1, Approach B). More recently, fusion proteins have been prepared of streptavidin with different enzymes and native fluorescent proteins.

In many cases, such as affinity chromatographic applications, more reversible binding of biotin to avidin would be a distinct advantage. In this context, streptavidin has a critical disadvantage in that the interaction between its subunits is very strong, and it cannot be used when reversibility of the interaction is desired. This is in contrast to avidin, from which an immobilized monovalent form can be produced (8), and the immobilized avidin monomer binds biotinylated compounds reversibly (9).

Site-directed mutagenesis and chemical modification studies are currently being performed on both avidin and streptavidin to understand better their interaction with biotin, the interaction between their subunits, and, perhaps eventually, for better application in the previously-mentioned systems (10 ,11)In this regard, the single, critical tyrosine residue of the binding sites of avidin and streptavidin was nitrated, and the tetrameric structures of the resultant “nitro avidin” and “nitro streptavidinwere retained (12). The biotin-binding property also had sufficiently high affinity for a variety of applications, including affinity chromatography, enzyme immobilization, and phage-display technology (13, 14). The major difference between the nitro avidins and the native molecules is that biotinylated compounds are released by competition with free biotin, and then the latter is liberated by treating the column with basic solutions (pH 10), thereby regenerating the original biotin-binding capacity of the nitro avidin affinity column. Reduced or altered binding characteristics are also conferred on the binding sites of avidin or streptavidin by site-directed mutagenesis of selected binding site residues, such as tryptophans.

References

1. M. Wilchek and E. A. Bayer, eds. (1990) Avidin-Biotin Technology Methods in Enzymology, Vol. 184, Academic Press, San Diego

2. E. A. Bayer and M. Wilchek (1978) Trends Biochem. Sci. 3, N237–N239

3. E. A. Bayer and M. Wilchek (1980) Methods Biochem. Anal. 26, 1–45.

4. M. Wilchek and E. A. Bayer (1984) Immunol. Today 5, 39–43

5. M. Wilchek and E. A. Bayer (1989) In Protein Recognition of Immobilized Ligands (T. W. Hutchens, ed.), Alan R. Liss, New York, pp. 83–90

6. E. A. Bayer and M. Wilchek (1996) In Immunoassay (E. P. Diamandis and T. K. Christopoulos, eds.), Academic Press, San Diego, pp. 237–267.  7. G. Paganelli, P. Magnani, A. G. Siccardi, and F. Fazio (1995) In Cancer Therapy with Radiolabeled Antibodies (D. M. Goldenberg, ed.), CRC Press, Boca Raton, FL, pp. 239–254.

8. N. M. Green and E. J. Toms (1973) Biochem. J. 133, 687–700

9. K. P. Henrikson, S. H. G. Allen, and W. L. Maloy (1979) Anal. Biochem. 94, 366–370

10. A. Chilkoti, P. H. Tan, and P. S. Stayton (1995) Proc. Natl. Acad. Sci. USA 92, 1754–1758

11. A. Chilkoti, B. L. Schwartz, R. D. Smith, C. J. Long, and P. S. Stayton (1995) Bio/Technology 13, 1198-1204 .

12. E. Morag, E. A. Bayer, and M. Wilchek (1996) Biochem. J. 316, 193–199

13. E. Morag, E. A. Bayer, and M. Wilchek (1996) Anal. Biochem. 243, 257–263

14. M. E. M. Balass, E. A. Bayer, S. Fuchs, M. Wilchek, and E. Katchalski-Katzir (1996) Anal. Biochem. 243, 264–269. 




علم الأحياء المجهرية هو العلم الذي يختص بدراسة الأحياء الدقيقة من حيث الحجم والتي لا يمكن مشاهدتها بالعين المجرَّدة. اذ يتعامل مع الأشكال المجهرية من حيث طرق تكاثرها، ووظائف أجزائها ومكوناتها المختلفة، دورها في الطبيعة، والعلاقة المفيدة أو الضارة مع الكائنات الحية - ومنها الإنسان بشكل خاص - كما يدرس استعمالات هذه الكائنات في الصناعة والعلم. وتنقسم هذه الكائنات الدقيقة إلى: بكتيريا وفيروسات وفطريات وطفيليات.



يقوم علم الأحياء الجزيئي بدراسة الأحياء على المستوى الجزيئي، لذلك فهو يتداخل مع كلا من علم الأحياء والكيمياء وبشكل خاص مع علم الكيمياء الحيوية وعلم الوراثة في عدة مناطق وتخصصات. يهتم علم الاحياء الجزيئي بدراسة مختلف العلاقات المتبادلة بين كافة الأنظمة الخلوية وبخاصة العلاقات بين الدنا (DNA) والرنا (RNA) وعملية تصنيع البروتينات إضافة إلى آليات تنظيم هذه العملية وكافة العمليات الحيوية.



علم الوراثة هو أحد فروع علوم الحياة الحديثة الذي يبحث في أسباب التشابه والاختلاف في صفات الأجيال المتعاقبة من الأفراد التي ترتبط فيما بينها بصلة عضوية معينة كما يبحث فيما يؤدي اليه تلك الأسباب من نتائج مع إعطاء تفسير للمسببات ونتائجها. وعلى هذا الأساس فإن دراسة هذا العلم تتطلب الماماً واسعاً وقاعدة راسخة عميقة في شتى مجالات علوم الحياة كعلم الخلية وعلم الهيأة وعلم الأجنة وعلم البيئة والتصنيف والزراعة والطب وعلم البكتريا.