SGFP2熒光蛋白是一種綠色熒光蛋白變體，發(fā)表于2007年，來(lái)源于維多利亞水母。SGFP2綠色熒光蛋白激發(fā)光波長(zhǎng)495nm，發(fā)射光波長(zhǎng)512nm，可以用藍(lán)光激發(fā)。SGFP2綠色熒光蛋白具有中度酸敏感性，大小為26.9kDa。

SGFP2綠色熒光蛋白的研究與發(fā)現(xiàn)

熒光蛋白已成為細(xì)胞生物學(xué)中非常寶貴的工具。綠色熒光蛋白變體EGFP的應(yīng)用尤其廣泛。然而，包括EGFP在內(nèi)的熒光蛋白的使用可能會(huì)受到蛋白質(zhì)折疊效率低下的阻礙，導(dǎo)致蛋白質(zhì)聚集和熒光減少。這在原核細(xì)胞中尤其明顯。此外，增強(qiáng)型綠色熒光蛋白EGFP的藍(lán)色熒光蛋白變體EBFP因其微弱的熒光和快速的光漂白而很少使用。因此，努力提高蛋白質(zhì)折疊、熒光亮度和光穩(wěn)定性等性能非常重要。

2007年，阿姆斯特丹大學(xué)和荷蘭烏特勒支大學(xué)的科研人員發(fā)表文獻(xiàn)，關(guān)于改進(jìn)綠色熒光蛋白和藍(lán)色熒光蛋白在細(xì)菌和哺乳動(dòng)物細(xì)胞中的表達(dá)?？蒲腥藛T分別基于EGFP和EBFP創(chuàng)建了強(qiáng)增強(qiáng)綠色熒光蛋白（SGFP2）和強(qiáng)增強(qiáng)藍(lán)色熒光蛋白（SBFP2）。科研人員使用定點(diǎn)突變引入了幾種突變，這些突變被證明可以改善熒光蛋白EYFP和ECFP。

熒光蛋白SGFP2和SBFP2在體外表現(xiàn)出更快、更有效的蛋白質(zhì)折疊和加速的發(fā)色團(tuán)氧化。對(duì)于這兩種強(qiáng)烈增強(qiáng)的熒光蛋白，其光穩(wěn)定性提高了2倍，SBFP2的量子產(chǎn)率提高了3倍。提高的折疊效率降低了大腸桿菌中蛋白質(zhì)聚集的程度，從而使表達(dá)SGFP2的細(xì)菌的亮度比表達(dá)EGFP的細(xì)菌的透明度提高了7倍。表達(dá)SBFP2的細(xì)菌比表達(dá)EBFP的細(xì)菌熒光強(qiáng)度高16倍。在哺乳動(dòng)物細(xì)胞中，這種改善不太明顯。表達(dá)SGFP2的細(xì)胞比表達(dá)EGFP的細(xì)胞亮1.7倍，這顯然是由于更有效的蛋白質(zhì)表達(dá)和/或發(fā)色團(tuán)成熟。表達(dá)SBFP2的哺乳動(dòng)物細(xì)胞比表達(dá)EBFP的細(xì)胞亮3.7倍。這種亮度的增加與純化重組蛋白觀察到的固有亮度的增加非常相似。熒光蛋白SGFP2和SBFP2的成熟效率和光穩(wěn)定性的提高有助于檢測(cè)并延長(zhǎng)熒光成像的更大持續(xù)時(shí)間。

SGFP2綠色熒光蛋白的激發(fā)光和發(fā)射光

SGFP2綠色熒光蛋白激發(fā)光波長(zhǎng)495nm，發(fā)射光波長(zhǎng)512nm，可以用熒光手電筒GFPfinder-2101RB激發(fā)。

熒光手電筒GFPfinder-2101RB提供高強(qiáng)度藍(lán)色激發(fā)光，可穿透培養(yǎng)皿，激發(fā)愈傷組織、菌落、細(xì)胞、植物、動(dòng)物中表達(dá)的綠色熒光蛋白。GFP綠色熒光蛋白和EGFP增強(qiáng)型綠色熒光蛋白，都可以使用熒光手電筒進(jìn)行快速檢測(cè)。通過特制濾光片，可用肉眼觀察綠色熒光，并拍攝清晰的熒光照片。

熒光手電筒GFPfinder-2101RB檢測(cè)效率高、使用方便，價(jià)格僅為熒光顯微鏡的十分之一，是檢測(cè)綠色熒光蛋白的理想設(shè)備，得到眾多高校和科研機(jī)構(gòu)的好評(píng)和認(rèn)可！

下圖：熒光手電筒GFPfinder-2101RB

SGFP2 Green Fluorescent Protein

Fluorescent proteins have become an invaluable tool in cell biology. The green fluorescent protein variant EGFP is especially widely applied. Use of fluorescent proteins, including EGFP, however can be hindered by inefficient protein folding, resulting in protein aggregation and reduced fluorescence. This is especially profound in prokaryotic cells. Furthermore, EBFP, a blue fluorescent protein variant of EGFP, is rarely used because of its dim fluorescence and fast photobleaching. Thus, efforts to improve properties such as protein folding, fluorescence brightness, and photostability are important. Strongly enhanced green fluorescent proteins (SGFP2) and strongly enhanced blue fluorescent proteins (SBFP2) were created, based on EGFP and EBFP, respectively. We used site-directed mutagenesis to introduce several mutations, which were recently shown to improve the fluorescent proteins EYFP and ECFP. Fluorescent proteins SGFP2 and SBFP2 exhibit faster and more efficient protein folding and accelerated chromophore oxidation in vitro. For both strongly enhanced fluorescent proteins, the photostability was improved 2-fold and the quantum yield of SBFP2 was increased 3-fold. The improved folding efficiency reduced the extent of protein aggregation in Escherichia coli, thereby increasing the brightness of bacteria expressing SGFP2 7-fold compared to the brightness of those expressing EGFP. Bacteria expressing SBFP2 were 16-fold more fluorescent than those expressing EBFP. In mammalian cells, the improvements were less pronounced. Cells expressing SGFP2 were 1.7-fold brighter than those expressing EGFP, which was apparently due to more efficient protein expression and/or chromophore maturation. Mammalian cells expressing SBFP2 were 3.7-fold brighter than cells expressing EBFP. This increase in brightness closely resembled the increase in intrinsic brightness observed for the purified recombinant protein. The increased maturation efficiency and photostability of SGFP2 and SBFP2 fluorescent proteins facilitate detection and extend the maximum duration of fluorescence imaging.

Reference:

Improved Green and Blue Fluorescent Proteins for Expression in Bacteria and Mammalian Cells

Kremers G-J, Goedhart J, Van Den Heuvel Dj, Gerritsen Hc, Gadella Twj (2007). Biochemistry, 46(12) , 3775-3783. doi: 10.1021/bi0622874.