Green fluorescent protein TurboGFP

- Bright green fluorescence
- Fast maturation at a wide range of temperatures
- High pH-stability and photostability
- Proven suitability to generate stably transfected cell lines
- Destabilized variant is available
- Recommended for gene expression analysis and cell and organelle labeling

TurboGFP is an improved variant of the green fluorescent protein CopGFP cloned from copepod Pontellina plumata (Arthropoda; Crustacea; Maxillopoda; Copepoda) [Shagin et al., 2004]. It possesses bright green fluorescence (excitation/ emission max = 482/ 502 nm) that is visible earlier than fluorescence of other green fluorescent proteins. TurboGFP is mainly intended for applications where fast appearance of bright fluorescence is crucial. It is specially recommended for cell and organelle labeling and tracking the promoter activity. Destabilized TurboGFP variant allows accurate analysis of rapid and/or transient events in gene regulation.

Main properties

TurboGFP normalized excitation (thin line) and emission (thick line) spectra. Spectra viewer tool Download TurboGFP spectra (xls)

CHARACTERISTIC * Brightness is a product of extinction coefficient and quantum yield, divided by 1000. Molecular weight, kDa 26 Polypeptide length, aa 232 Fluorescence color green Excitation maximum, nm 482 Emission maximum, nm 502 Quantum yield 0.53 Extinction coefficient, M-1cm-1 70 000 Brightness* 37.1 Brightness, % of EGFP 112 pKa 5.2 Structure dimer Aggregation no Maturation rate at 37°C super fast Photostability high Cell toxicity not observed Main advantages superbright and fast-maturing green fluorescent protein Possible limitations dimer, limited applicability for fusions generation

Recommended filter sets and antibodies

TurboGFP can be recognized using Anti-TurboGFP(d) (Cat.# AB513) antibody available from Evrogen.

TurboGFP can be detected using common fluorescence filter sets for EGFP, FITC, and other green dyes. Recommended Omega Optical filter sets are QMAX-Green, XF100-2, XF100-3, XF115-2, and XF116-2.

Performance and use

TurboGFP can be expressed and detected in a wide range of organisms including cold-blooded animals. Mammalian cells transiently transfected with TurboGFP expression vectors produce bright fluorescence in 8-10 hrs after transfection. No cytotoxic effects or visible protein aggregation are observed. TurboGFP can be used in multicolor labeling applications with blue, true-yellow, red, and far-red fluorescent dyes.

	TurboGFP expression in transiently transfected mammalian cells.

TurboGFP suitability to generate stably transfected cells has been proven by Marinpharm company.

Despite its dimeric structure, TurboGFP performs well in some fusions. However, for protein labeling applications we recommend using specially optimized monomeric TagFPs.

TurboGFP expression in stably transfected mammalian cell lines. (A) Human cervix carcinoma HeLa cells with TurboGFP in cytoplasm; (B) C2C12 mouse myoblasts with TurboGFP in cytoplasm; (C) PC-12 rat phaeochromocytoma cells with TurboGFP in cytoplasm; (D) PC-12 rat phaeochromocytoma cells with TurboGFP in cytoplasm after the addition of nerve growth factor (cells differentiate irreversibly into neuron-like cells); (G) Walker 256 rat tumour cells with TurboGFP in cytoplasm; (H) M3-mouse melanoma with TurboGFP in cytoplasm; (I) 3T3 mouse fibroblast with TurboGFP in cytoplasm; (J) T24 human bladder carcinoma with TurboGFP in cytoplasm; (E) Chinese Hamster Ovary Cells CHO-K1 with TurboGFP in cytoplasm; (F) HeLa cells expressing TurboGFP-fibrillarin fusion; (K) HeLa cells expressing mitochondria-targeted TurboGFP; (L) T24 human bladder carcinoma expressing mitochondria-targeted TurboGFP. Images were kindly provided by Dr. Christian Petzelt (Marinpharm).

TurboGFP maturation kinetics: TurboGFP allows monitoring the activity from early promoters. It matures noticeably faster than EGFP and most other fluorescent proteins. This difference in performance is illustrated here using both in vitro analysis of TurboGFP and EGFP refolding and maturation kinetics and in vivo examination of the developing Xenopus embrios expressing either TurboGFP or EGFP.

		Comparison of EGFP (violet lines) and TurboGFP (green lines) refolding and maturation speed in vitro. Normalized fluorescence recovery plots are shown. (A) — refolding kinetics; (B) — chromophore maturation kinetics.

		In vivo comparison of TurboGFP and EGFP maturation in developing Xenopus embryos Vectors encoding TurboGFP and EGFP under the control of CMV promoter were microinjected into animal poles of Xenopus embryos at the stage of two blastomeres. Living embryos were then photographed from the animal pole at the middle and late gastrula stages. Experimental data were presented by Dr. A. Zaraisky, Institute of Bioorganic Chemistry, RAS (Moscow, Russia).

Available variants and fusions

Variant	Description	Related vector	Cat.#	Click for image
Humanized TurboGFP	TurboGFP codon usage is optimized for high expression in mammalian cells [Haas et al., 1996], but it can be successfully expressed in many other heterological systems.	pTurboGFP-C	FP511
		pTurboGFP-N	FP512
		pTurboGFP-B	FP513
		pTurboGFP-PRL	FP515
		Gateway® TurboGFP-C entry clone	FP521
		Gateway® TurboGFP-N entry clone	FP522
Destabilized TurboGFP variant (TurboGFP-dest1)	TurboGFP-dest1 is produced by fusing the initial protein with PEST amino acid sequence encoded by region 422-461 of mouse ornithine decarboxylase gene [Li et al., 1998]. This sequence targets the protein to degradation and enables a rapid protein turnover. TurboGFP-dest1 retains spectral properties of the initial protein, but has shorter half-life (approximately 2 hrs) as measured by the analysis of fluorescence intensity of cells treated with a protein synthesis inhibitor, cycloheximide. Because of rapid turnover, TurboGFP-dest1 can be used to measure changes in gene expression.	peTurboGFP-PRL-dest1	FP523
		peTurboGFP-dest1	FP524
TurboGFP-mito fusion	A mitochondrial targeting sequence (MTS) is fused to the TurboGFP N-terminus. MTS was derived from the subunit VIII of human cytochrome C oxidase [Rizzuto et al., 1989; Rizzuto et al., 1995]. When expressed in mammalian cells, this variant provides green fluorescent labeling of mitochondria.	pTurboGFP-mito	FP517