Baculovirus-infected insect cells have become a powerful tool to express recombinant proteins for structural and functional studies by NMR spectroscopy. This article provides an introduction into the insect cell/baculovirus expression system and its use for the production of recombinant isotope-labeled proteins. We discuss recent advances in inexpensive isotope-labeling methods using labeled algal or yeast extracts as the amino acid source and give examples of advanced NMR applications for proteins, which have become accessible by this eukaryotic expression host.
Application Note: Efficient Uniform Labeling of Proteins Expressed in Baculovirus-Infected Insect Cells Using ... - SelectScience
Application Note: Efficient Uniform Labeling of Proteins Expressed in Baculovirus-Infected Insect Cells Using ... - SelectScience
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Application Note: Efficient Uniform Labeling of Proteins Expressed in Baculovirus-Infected Insect Cells Using ...
SelectScience
Uniform isotope labeling is a key tool for NMR studies on recombinant proteins and their interaction with ligands of pharmaceutical interest. To be useful for most important NMR studies, a labeled protein has to be expressed with baculovirus-infected ...
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07-23-2017 11:38 AM
An improved protocol for amino acid type-selective isotope labeling in insect cells
An improved protocol for amino acid type-selective isotope labeling in insect cells
Abstract
An improved expression protocol is proposed for amino acid type-specific , -isotope labeling of proteins in baculovirus-infected (BV) insect cell cultures. This new protocol modifies the methods published by Gossert et al. (J Biomol NMR 51(4):449â??456, 2011) and provides efficient incorporation of isotopically labeled amino acids, with similar yields per L versus unlabeled expression in rich media. Gossert et al. identified the presence of unlabeled amino...
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07-15-2017 05:05 PM
An economic approach to efficient isotope labeling in insect cells using homemade 15 N-, 13 C- and 2 H-labeled yeast extracts
An economic approach to efficient isotope labeling in insect cells using homemade 15 N-, 13 C- and 2 H-labeled yeast extracts
Abstract
Heterologous expression of proteins in insect cells is frequently used for crystallographic structural studies due to the high yields even for challenging proteins requiring the eukaryotic protein processing capabilities of the host. However for NMR studies, the need for isotope labeling poses extreme challenges in eukaryotic hosts. Here, we describe a robust method to achieve uniform protein 15N and 13C labeling of up...
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06-12-2015 07:48 PM
Affordable uniform isotope labeling with 2 H, 13 C and 15 N in insect cells
Affordable uniform isotope labeling with 2 H, 13 C and 15 N in insect cells
Abstract
For a wide range of proteins of high interest, the major obstacle for NMR studies is the lack of an affordable eukaryotic expression system for isotope labeling. Here, a simple and affordable protocol is presented to produce uniform labeled proteins in the most prevalent eukaryotic expression system for structural biology, namely Spodoptera frugiperda insect cells. Incorporation levels of 80Â*% can be achieved for 15N and 13C with yields comparable to expression in...
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04-30-2015 09:13 PM
[NMR paper] High-Resolution Heteronuclear Multidimensional NMR of Proteins in Living Insect Cells Using a Baculovirus Protein Expression System.
High-Resolution Heteronuclear Multidimensional NMR of Proteins in Living Insect Cells Using a Baculovirus Protein Expression System.
http://www.ncbi.nlm.nih.gov/corehtml/query/egifs/http:--pubs.acs.org-images-pubmed-acspubs.jpg Related Articles High-Resolution Heteronuclear Multidimensional NMR of Proteins in Living Insect Cells Using a Baculovirus Protein Expression System.
J Am Chem Soc. 2013 Jan 27;
Authors: Hamatsu J, O'Donovan D, Tanaka T, Shirai T, Hourai Y, Mikawa T, Ikeya T, Mishima M, Boucher W, Smith BO, Laue ED, Shirakawa M, Ito Y
...
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02-03-2013 10:19 AM
High-Resolution HeteronuclearMultidimensional NMRof Proteins in Living Insect Cells Using a Baculovirus Protein ExpressionSystem
High-Resolution HeteronuclearMultidimensional NMRof Proteins in Living Insect Cells Using a Baculovirus Protein ExpressionSystem
Jumpei Hamatsu, Daniel O’Donovan, Takashi Tanaka, Takahiro Shirai, Yuichiro Hourai, Tsutomu Mikawa, Teppei Ikeya, Masaki Mishima, Wayne Boucher, Brian O. Smith, Ernest D. Laue, Masahiro Shirakawa and Yutaka Ito
http://pubs.acs.org/appl/literatum/publisher/achs/journals/content/jacsat/0/jacsat.ahead-of-print/ja310928u/aop/images/medium/ja-2012-10928u_0005.gif
Journal of the American Chemical Society
DOI: 10.1021/ja310928u...
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01-27-2013 11:41 PM
A simple protocol for amino acid type selective isotope labeling in insect cells with improved yields and high reproducibility
A simple protocol for amino acid type selective isotope labeling in insect cells with improved yields and high reproducibility
Abstract An easy to use and robust approach for amino acid type selective isotope labeling in insect cells is presented. It relies on inexpensive commercial media and can be implemented in laboratories without sophisticated infrastructure. In contrast to previous protocols, where either high protein amounts or high incorporation ratios were obtained, here we achieve both at the same time. By supplementing media with a well considered amount of yeast extract,...
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10-05-2011 08:57 PM
Production of isotopically labeled heterologous proteins in non-E. coli prokaryotic and eukaryotic cells
Production of isotopically labeled heterologous proteins in non-E. coli prokaryotic and eukaryotic cells
Abstract The preparation of stable isotope-labeled proteins is necessary for the application of a wide variety of NMR methods, to study the structures and dynamics of proteins and protein complexes. The E. coli expression system is generally used for the production of isotope-labeled proteins, because of the advantages of ease of handling, rapid growth, high-level protein production, and low cost for isotope-labeling. However, many eukaryotic proteins are not functionally expressed...
Production of isotope-labeled proteins in insect cells for NMR
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