Archaeal chromatin proteins Cren7 and Sul7d from are DNA benders. To better understand their architectural roles in chromosomal DNA organization, we analyzed DNA compaction by Cren7 and Sis7d, a Sul7d family member, from at the single-molecule (SM) level by total single-molecule internal reflection fluorescence microscopy (SM-TIRFM) and atomic force microscopy (AFM). We show that both Cren7 and Sis7d were able to compact singly tethered λ DNA into a highly condensed structure in a three-step process and that Cren7 was over an order of magnitude more efficient than Sis7d in DNA compaction.
View Article and Find Full Text PDFNucleoid-associated proteins (NAPs) play an important role on chromosome condensation and organization. Mycobacterial integration host factor (mIHF) is one of the few mycobacterial NAPs identified so far. mIHF has the ability to stimulate mycobacteriophage L5 integration and compact DNA into nucleoid-like or higher order filamentous structures by atomic force microscopy observation.
View Article and Find Full Text PDFThe fluorescence microscopy has made great contributions in dissecting the mechanisms of complex biological processes at the single molecule level. In single molecule assays for studying DNA-protein interactions, there are two important factors for consideration: the DNA substrate with enough length for easy observation and labeling a protein with a suitable fluorescent probe. 48.
View Article and Find Full Text PDFMuch of our knowledge in conventional biochemistry has derived from bulk assays. However, many stochastic processes and transient intermediates are hidden when averaged over the ensemble. The powerful technique of single-molecule fluorescence microscopy has made great contributions to the understanding of life processes that are inaccessible when using traditional approaches.
View Article and Find Full Text PDFUnlabelled: SSV-type integrases, encoded by fuselloviruses which infect the hyperthermophilic archaea of the Sulfolobales, are archaeal members of the tyrosine recombinase family. These integrases catalyze viral integration into and excision from a specific site on the host genome. In the present study, we have established an in vitro integration/excision assay for SSV2 integrase (Int(SSV2)).
View Article and Find Full Text PDFActa Crystallogr D Biol Crystallogr
June 2012
The spindle-shaped virus SSV1 of the hyperthermophilic archaeon Sulfolobus shibatae encodes an integrase (SSV1 Int). Here, the crystal structure of the C-terminal catalytic domain of SSV1 Int is reported. This is the first structural study of an archaeal tyrosine recombinase.
View Article and Find Full Text PDFThe accuracy of DNA synthesis by DNA polymerase B1 from the hyperthermophilic archaeon Sulfolobus solfataricus (Sso pol B1) at near the physiological temperature was investigated using M13-based mutational assays. Sso pol B1 showed replication fidelity similar to or higher than most viral, bacterial, and eukaryotic replicases. The fidelity of the enzyme was about three times as high at 70 degrees C as at 55 degrees C.
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