分类: 生物学 >> 生物物理学 提交时间: 2016-05-11
摘要: Lysine acetylation is an important post-translational modification and is known to regulate many eukaryotic cellular processes. Little, however, is known about acetylated proteins in prokaryotes. Here, using immunoblotting, mass spectrometry and mutagenesis studies, we investigate the acetylation dynamics of the DNA repair protein Ku and its relationship with the deacetylase protein Sir2 and the non-homologous end joining (NHEJ) pathway in Mycobacterium smegmatis. We report that acetylation of Ku increases with growth, while NHEJ activity decreases, providing support for the hypothesis that acetylation of Ku may be involved in the DNA damage response in bacteria. Ku has multiple lysine sites. Our results indicate that K29 is an important acetylation site and that deficiency of Sir2 or mutation of K29 affects the quantity of Ku and its acetylation dynamics. Our findings expand knowledge of acetylation targets in prokaryotes and indicate a new direction for further research on bacterial DNA repair mechanisms.
分类: 生物学 >> 生物物理学 提交时间: 2016-05-11
摘要: Electron transfer (ET) is widely used for driving the processes that underlie the chemistry of life. However, our abilities to probe electron transfer mechanisms in proteins and design redox enzymes are limited, due to :the lack of methods to site-specifically insert electron acceptors into proteins in vivo. Here we describe the synthesis and genetic incorporation of 4-fluoro-3-nitrophenylalanine (FNO(2)Phe), which has similar reduction potentials to NAD(P)H and ferredoxin, the most important biological reductants. Through the genetic incorporation of FNO2Phe into green fluorescent:protein (GFP) and femtosecond transient absorption measurement, we show that photoinduced electron transfer (PET) from the GFP chromophore to FNO2Phe occurs very fast (within 11 ps), which is comparable to that of the first electron transfer step in photosystem I, from P700* to A(0). This genetically encoded, low-reduction potential unnatural amino acid (UAA) can significantly in-Trove our ability to investigate electron transfer mechanisms in complex reductases and facilitate the design of miniature proteins that mimic their functions.