The Experts below are selected from a list of 24 Experts worldwide ranked by ideXlab platform
Rosaria P Haugland - One of the best experts on this subject based on the ideXlab platform.
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easily reversible desthiobiotin binding to streptavidin avidin and other biotin binding Proteins uses for protein labeling detection and isolation
Analytical Biochemistry, 2002Co-Authors: James Hirsch, Leila Eslamizar, Brian Filanoski, Nabi Malekzadeh, Rosaria P Haugland, Joseph M BeechemAbstract:The high-affinity binding of biotin to avidin, streptavidin, and related Proteins has been exploited for decades. However, a disadvantage of the biotin/Biotin-Binding protein interaction is that it is essentially irreversible under physiological conditions. Desthiobiotin is a biotin analogue that binds less tightly to Biotin-Binding Proteins and is easily displaced by biotin. We synthesized an amine-reactive desthiobiotin derivative for labeling Proteins and a desthiobiotin-agarose affinity matrix. Conjugates labeled with desthiobiotin are equivalent to their biotinylated counterparts in cell-staining and antigen-labeling applications. They also bind to streptavidin and other Biotin-Binding protein-based affinity columns and are recognized by anti-biotin antibodies. Fluorescent streptavidin conjugates saturated with desthiobiotin, but not biotin, bind to a cell-bound biotinylated target without further processing. Streptavidin-based ligands can be gently stripped from desthiobiotin-labeled targets with buffered biotin solutions. Thus, repeated probing with fluorescent streptavidin conjugates followed by enzyme-based detection is possible. In all applications, the desthiobiotin/Biotin-Binding protein complex is easily dissociated under physiological conditions by either biotin or desthiobiotin. Thus, our desthiobiotin-based reagents and techniques provide some distinct advantages over traditional 2-iminobiotin, monomeric avidin, or other affinity-based techniques.
James Hirsch - One of the best experts on this subject based on the ideXlab platform.
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easily reversible desthiobiotin binding to streptavidin avidin and other biotin binding Proteins uses for protein labeling detection and isolation
Analytical Biochemistry, 2002Co-Authors: James Hirsch, Leila Eslamizar, Brian Filanoski, Nabi Malekzadeh, Rosaria P Haugland, Joseph M BeechemAbstract:The high-affinity binding of biotin to avidin, streptavidin, and related Proteins has been exploited for decades. However, a disadvantage of the biotin/Biotin-Binding protein interaction is that it is essentially irreversible under physiological conditions. Desthiobiotin is a biotin analogue that binds less tightly to Biotin-Binding Proteins and is easily displaced by biotin. We synthesized an amine-reactive desthiobiotin derivative for labeling Proteins and a desthiobiotin-agarose affinity matrix. Conjugates labeled with desthiobiotin are equivalent to their biotinylated counterparts in cell-staining and antigen-labeling applications. They also bind to streptavidin and other Biotin-Binding protein-based affinity columns and are recognized by anti-biotin antibodies. Fluorescent streptavidin conjugates saturated with desthiobiotin, but not biotin, bind to a cell-bound biotinylated target without further processing. Streptavidin-based ligands can be gently stripped from desthiobiotin-labeled targets with buffered biotin solutions. Thus, repeated probing with fluorescent streptavidin conjugates followed by enzyme-based detection is possible. In all applications, the desthiobiotin/Biotin-Binding protein complex is easily dissociated under physiological conditions by either biotin or desthiobiotin. Thus, our desthiobiotin-based reagents and techniques provide some distinct advantages over traditional 2-iminobiotin, monomeric avidin, or other affinity-based techniques.
Takeshi Yamamoto - One of the best experts on this subject based on the ideXlab platform.
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tamavidin a versatile affinity tag for protein purification and immobilization
Journal of Biotechnology, 2010Co-Authors: Yoshimitsu Takakura, Masako Tsunashima, Satoru Usami, Naomi Oka, Hitomi Kajiwara, Hiroshi Tsukamoto, Yuji Ishida, Takeshi YamamotoAbstract:Abstract Tamavidin 2 is a fungal avidin-like protein that binds biotin with high affinity and is highly produced in soluble form in Escherichia coli. By contrast, widely used Biotin-Binding Proteins avidin and streptavidin are rarely produced in soluble form in E. coli. In this study, we describe an efficient system for one-step purification and immobilization of recombinant Proteins using tamavidin 2 as an affinity tag. A bacterial sialyltransferase and soybean agglutinin were fused to tamavidin 2 and expressed in E. coli and tobacco BY-2 cells, respectively. High-level expressions of the fusion Proteins were detected (80 mg l−1 E. coli culture for bacterial sialyltransferase–tamavidin 2 and 2 mg l−1 BY-2 cell culture for soybean agglutinin–tamavidin 2). To immobilize and purify the fusion Proteins, biotinylated magnetic microbeads were incubated with the soluble extract from each recombinant host producing the fusion protein and then washed thoroughly. As the result, both fusion Proteins were immobilized tightly on the microbeads without substantial loss of activity and simultaneously highly purified (90–95% purity) on the microbeads. Biotin with a longer linker contributed to higher affinity between the fusion protein and biotin. These results suggest that tamavidin fusion technology is a powerful tool for production, purification, and immobilization of recombinant Proteins.
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tamavidins novel avidin like biotin binding Proteins from the tamogitake mushroom
FEBS Journal, 2009Co-Authors: Yoshimitsu Takakura, Masako Tsunashima, Junko Suzuki, Satoru Usami, Yoshimitsu Kakuta, Nozomu Okino, Makoto Ito, Takeshi YamamotoAbstract:Novel Biotin-Binding Proteins, referred to herein as tamavidin 1 and tamavidin 2, were found in a basidiomycete fungus, Pleurotus cornucopiae, known as the Tamogitake mushroom. These are the first avidin-like Proteins to be discovered in organisms other than birds and bacteria. Tamavidin 1 and tamavidin 2 have amino acid sequences with 31% and 36% identity, respectively, to avidin, and 47% and 48% identity, respectively, to streptavidin. Unlike any other Biotin-Binding Proteins, tamavidin 1 and tamavidin 2 are expressed as soluble Proteins at a high level in Escherichia coli. Recombinant tamavidin 2 was purified as a tetrameric protein in a single step by 2-iminobiotin affinity chromatography, with a yield of 5 mg per 100 mL culture of E. coli. The kinetic parameters measured by a BIAcore biosensor indicated that recombinant tamavidin 2 binds biotin with high affinity, in a similar manner to binding by avidin and streptavidin. The overall crystal structure of recombinant tamavidin 2 is similar to that of avidin and streptavidin. However, recombinant tamavidin 2 is immunologically distinct from avidin and streptavidin. Tamavidin 2 and streptavidin are very similar in terms of the arrangement of the residues interacting with biotin, but different with regard to the number of hydrogen bonds to biotin carboxylate. Recombinant tamavidin 2 is more stable than avidin and streptavidin at high temperature, and nonspecific binding to DNA and human serum by recombinant tamavidin 2 is lower than that for avidin. These findings highlight tamavidin 2 as a probable powerful tool, in addition to avidin and streptavidin, in numerous applications of Biotin-Binding Proteins.
Brian Filanoski - One of the best experts on this subject based on the ideXlab platform.
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easily reversible desthiobiotin binding to streptavidin avidin and other biotin binding Proteins uses for protein labeling detection and isolation
Analytical Biochemistry, 2002Co-Authors: James Hirsch, Leila Eslamizar, Brian Filanoski, Nabi Malekzadeh, Rosaria P Haugland, Joseph M BeechemAbstract:The high-affinity binding of biotin to avidin, streptavidin, and related Proteins has been exploited for decades. However, a disadvantage of the biotin/Biotin-Binding protein interaction is that it is essentially irreversible under physiological conditions. Desthiobiotin is a biotin analogue that binds less tightly to Biotin-Binding Proteins and is easily displaced by biotin. We synthesized an amine-reactive desthiobiotin derivative for labeling Proteins and a desthiobiotin-agarose affinity matrix. Conjugates labeled with desthiobiotin are equivalent to their biotinylated counterparts in cell-staining and antigen-labeling applications. They also bind to streptavidin and other Biotin-Binding protein-based affinity columns and are recognized by anti-biotin antibodies. Fluorescent streptavidin conjugates saturated with desthiobiotin, but not biotin, bind to a cell-bound biotinylated target without further processing. Streptavidin-based ligands can be gently stripped from desthiobiotin-labeled targets with buffered biotin solutions. Thus, repeated probing with fluorescent streptavidin conjugates followed by enzyme-based detection is possible. In all applications, the desthiobiotin/Biotin-Binding protein complex is easily dissociated under physiological conditions by either biotin or desthiobiotin. Thus, our desthiobiotin-based reagents and techniques provide some distinct advantages over traditional 2-iminobiotin, monomeric avidin, or other affinity-based techniques.
Nabi Malekzadeh - One of the best experts on this subject based on the ideXlab platform.
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easily reversible desthiobiotin binding to streptavidin avidin and other biotin binding Proteins uses for protein labeling detection and isolation
Analytical Biochemistry, 2002Co-Authors: James Hirsch, Leila Eslamizar, Brian Filanoski, Nabi Malekzadeh, Rosaria P Haugland, Joseph M BeechemAbstract:The high-affinity binding of biotin to avidin, streptavidin, and related Proteins has been exploited for decades. However, a disadvantage of the biotin/Biotin-Binding protein interaction is that it is essentially irreversible under physiological conditions. Desthiobiotin is a biotin analogue that binds less tightly to Biotin-Binding Proteins and is easily displaced by biotin. We synthesized an amine-reactive desthiobiotin derivative for labeling Proteins and a desthiobiotin-agarose affinity matrix. Conjugates labeled with desthiobiotin are equivalent to their biotinylated counterparts in cell-staining and antigen-labeling applications. They also bind to streptavidin and other Biotin-Binding protein-based affinity columns and are recognized by anti-biotin antibodies. Fluorescent streptavidin conjugates saturated with desthiobiotin, but not biotin, bind to a cell-bound biotinylated target without further processing. Streptavidin-based ligands can be gently stripped from desthiobiotin-labeled targets with buffered biotin solutions. Thus, repeated probing with fluorescent streptavidin conjugates followed by enzyme-based detection is possible. In all applications, the desthiobiotin/Biotin-Binding protein complex is easily dissociated under physiological conditions by either biotin or desthiobiotin. Thus, our desthiobiotin-based reagents and techniques provide some distinct advantages over traditional 2-iminobiotin, monomeric avidin, or other affinity-based techniques.