Functional analysis of the domain organization of Trypanosoma brucei RNase HI.
The structure-function relationship of Trypanosoma brucei RNase HI was investigated by evaluating the abilities of truncated forms of the enzyme to convert RNase H substrate to product. Our studies identify a 42-amino-acid noncanonical RNase HI spacer domain essential for function. We also show that the enzyme's nuclear localization domain is not required for RNase H activity but functions as an RNA binding domain which modulates the enzyme's Mn(2+)-dependent activity. These findings show that the enzyme's RNA binding/nuclear targeting and RNase H activities are organized into discrete N- and C-terminal domains with boundaries established by its spacer domain. This is the first report of the unusual structure to function relationship of a protozoal RNase H. This relationship may be conserved in other eukaryotic RNases H suggesting that criteria preserving their structure and function may be important to their roles in nucleic acid metabolism.
Kobil
J H
JH
Department of Molecular Microbiology and Immunology, Brown University, Providence, Rhode Island 02912, USA.
Campbell
A G
AG
D000595:Amino Acid Sequence; D000818:Animals; D001483:Base Sequence; D002467:Cell Nucleus / Q000378:metabolism; D017931:DNA Primers; D006801:Humans; D008969:Molecular Sequence Data; D011485:Protein Binding; D012313:RNA / Q000378:metabolism; D016914:Ribonuclease H / Q000737:chemistry / Q000378:metabolism*; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship; D014346:Trypanosoma brucei brucei / Q000201:enzymology*
D017931:DNA Primers; D012313:RNA; D016914:Ribonuclease H; C078856:ribonuclease HI