Abstract
Sulfolobus solfataricus N-terminus and other regions of the partial amino acid sequence of a thermoprotein exhibiting poly(ADP-ribose) polymerase activity suggest that it belongs to the DINGGG class of proteins that are often described as membrane bound. Our previous biochemical studies demonstrated that the thermoprotein is also strictly associated with DNA, and is only partially solubilized from cell homogenate. The present research is focused on the analysis of the sulfolobal DING thermozyme localization within the archaeal cell. Immunofluorescence microscopy evidenced the peripheral cell localization of Sulfolobal DING protein, along the plasma membrane hedge. Less intense, but clearly occurring, is the merge of Sulfolobus poly (ADP-ribose) polymerase with nucleoid. Anti-poly(ADP-ribose) polymerase immunoblottings clearly showed the occurrence of Sulfolobus thermozyme in membrane fractions as well as they confirmed its association with nucleoid DNA. Fluorescent anti-PARP-1 antibodies showed that the PARPSso immunosignal localizes close to the membrane, at the periphery of cell, and that PARPSso green signal is also overlapping or strictly close to the nucleoid. Biochemical analyses confirmed that the thermozyme occurs in both membrane and nucleoid preparations.
Highlights
Sulfolobus solfataricus N-terminus and other regions of the partial amino acid sequence of a thermoprotein exhibiting poly(ADP-ribose) polymerase activity suggest that it belongs to the DINGGG class of proteins that are often described as membrane bound
Intracellular localization of PARPSso by fluorescence microscopy The intracellular localization of PARPSso and its relationship with cell components was investigated by fluorescence microscopy
The fluorescent anti-poly-ADP-ribose polymerase (PARP)-1 staining outlines the perimeter of the S. solfataricus cells (Fig. 1a, bottom left; green), as does the membrane staining (Fig. 1a, top, right; red), while the DAPI-stained DNA is located in the cell centre (Fig. 1a, top middle; blue)
Summary
Sulfolobus solfataricus N-terminus and other regions of the partial amino acid sequence of a thermoprotein exhibiting poly(ADP-ribose) polymerase activity suggest that it belongs to the DINGGG class of proteins that are often described as membrane bound. Our previous biochemical studies demonstrated that the thermoprotein is strictly associated with DNA, and is only partially solubilized from cell homogenate. The present research is focused on the analysis of the sulfolobal DING thermozyme localization within the archaeal cell. DING proteins (38–40 kDa) are ubiquitous, characterized by a conserved N-terminal sequence, DINGGG [1]. First identified in several animal and plant tissues, and in most eukaryotes and bacteria [2], at present the DING family includes about thirty members that are usually secreted, some are found in the cytosol [2, 3].
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