Structure-Based Genetic Analysis of Hel308a in the Hyperthermophilic Archaeon Sulfolobus islandicus.
In archaea, the HEL308 homolog Hel308a (or Hjm) is implicated in stalled replication fork repair. The biochemical properties and structures of Hjm homologs are well documented, but in vivo mechanistic information is limited. Herein, a structure-based functional analysis of Hjm was performed in the genetically tractable hyperthermophilic archaeon, Sulfolobus islandicus. Results showed that domain V and residues within it, which affect Hjm activity and regulation, are essential and that the domain V-truncated mutants and site-directed mutants within domain V cannot complement hjm chromosomal deletion. Chromosomal hjm deletion can be complemented by ectopic expression of hjm under the control of its native promoter but not an artificial arabinose promoter. Cellular Hjm levels are kept constant under ultraviolet (UV) and methyl methanesulfonate (MMS) treatment conditions in a strain carrying a plasmid to induce Hjm overexpression. These results suggest that Hjm expression and activity are tightly controlled, probably at the translational level.
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China.
Xueguo
X
Song
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China.
Jinfeng
J
Ni
State Key Laboratory of Microbial Technology, Shandong University, Jinan 250100, China. Electronic address: yulgshen@sdu.edu.cn.
Yulong
Y
Shen
D000595:Amino Acid Sequence; D019843:Archaeal Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D002470:Cell Survival; D008958:Models, Molecular; D011401:Promoter Regions, Genetic / Q000235:genetics; D000072417:Protein Domains; D016944:Sulfolobus / Q000166:cytology / Q000201:enzymology / Q000235:genetics* / Q000502:physiology; D013696:Temperature
D019843:Archaeal Proteins
Archaea; Genetic complementation; Hel308a; Replication fork regression; Sulfolobus islandicus