Newly isolated archaerhodopsin from a strain of Chinese halobacteria and its proton pumping behavior

Abstract

A strain of extremely salt-loving halobacteria Halobacterium species xz515 from a salt lake in Tibet was isolated. SDS-polyacrylamide gel electrophoresis shows that there is only one protein on claret membrane, which is the same membrane fraction as purple membrane from Halobacterium salinarum, with a molecular weight close to bacteriorhodopsin (br). The purified retinal containing protein from xz515 has an absorption peak at around 550 nm. These facts indicate that it is a br-like protein. The partial sequence determination [H. Wang et al., Chin. Sci. Bull., 45 (2000)] shows that this br-like protein belongs to the archaerhodopsin family. The measurements of light-induced medium pH change in intact cells and cell envelope vesicles of xz515 suggest that this type of archaerhodopsin has a proton pumping function. However, the study about the dynamics of pumped protons across the membrane reveal that the proton release and proton uptake is in reverse order compared to br. The probable reason, attributing to regulating the rate of proton release is discussed.

Novel expression and characterization of a light driven proton pump archaerhodopsin 4 in a Halobacterium salinarum strain

“…2C), similar to the retinal binding in BR [37]. However, the UV-VIS spectrum of AR4-xz515 shows a three peak pattern at 478 nm, 507 nm and 543 nm, respectively, which has been attributed to the presence of a C50-carotenoid lipid in the claret membrane, named as bacterioruberin [18]. Clearly, bacterioruberin is absent in the bAR membrane and the UV-VIS spectrum is similar to the bacterioruberin-excluded AR4-xz515, as reported before [18].…”

“…The proton pumping activities of the native and recombinant AR4 proteins were monitored through a light-induced absorption change by a pH-sensitive dye, pyranine (8-hydroxy-1,3,6-pyrenetrisulfonic acid, trisodium salt) on a homemade apparatus as described previously [18,23,39]. The net proton pumping activity was determined by taking the absorbance difference at 456 nm before and after adding the dye.…”

“…However, the UV-VIS spectrum of AR4-xz515 shows a three peak pattern at 478 nm, 507 nm and 543 nm, respectively, which has been attributed to the presence of a C50-carotenoid lipid in the claret membrane, named as bacterioruberin [18]. Clearly, bacterioruberin is absent in the bAR membrane and the UV-VIS spectrum is similar to the bacterioruberin-excluded AR4-xz515, as reported before [18]. A small blue shift in bAR when compared with BR may indicate that a slightly different protein packing, which may affect the retinal binding pocket in bAR [53], and hydrophobic binding pocket in rhodopsin has also been shown to be functionally important [54].…”

“…It serves as a model for investigating the vectorial ion translocation across membranes [8][9][10][11] and a template for G protein-coupled receptors [12], as it is a highly stable protein under experimental conditions [13,14]. Archaerhodopsins (ARs) are also light-driven proton pumps with a high degree of amino acid sequence homology to BR, and exhibit similar photochemical properties [15][16][17][18].…”

“…sp.) xz515, was collected from a salt lake in Tibet [18][19][20][21][22]. It was named as the fourth archaerhodopsin, as it exhibits 87% sequence similarity to AR1, 97% to AR2 and 84% to AR3.…”