Functional analysis of conserved residues in the putative "finger" domain of telomerase reverse transcriptase.
Telomerase is a ribonucleoprotein reverse transcriptase (RT) responsible for the maintenance of one strand of telomere terminal repeats. The catalytic protein subunit of telomerase, known generically as telomerase reverse transcriptase (TERT), exhibits significant homology to RTs encoded by retroviruses and retroelements. The polymerization mechanisms of telomerase may therefore be similar to those of the "conventional" RTs. In this study, we explored the extent of mechanistic conservation by analyzing mutations of conserved residues within the putative "finger" domain of TERT. Previous analysis has implicated this domain of retroviral RTs in nucleotide and RNA binding and in processivity control. Our results demonstrate that residues conserved between TERT and human immunodeficiency virus-1 RT are more likely than TERT-specific residues to be required for enzyme activity. In addition, residues presumed to make direct contact with either the RNA or nucleotide substrate appear to be functionally more important. Furthermore, distinct biochemical defects can be observed for alterations in the putative RNA- and nucleotide-binding TERT residues in a manner that can be rationalized by their postulated mechanisms of action. This study thus supports a high degree of mechanistic conservation between telomerase and retroviral RTs and underscores the roles of distinct aspects of telomerase biochemistry in telomere length maintenance.
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Department of Microbiology and Immunology, William Randolph Hearst Microbiology Research Center, Weill Medical College, Cornell University, New York, New York 10021, USA.
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D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001483:Base Sequence; D017931:DNA Primers; D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; D054303:HIV Reverse Transcriptase / Q000737:chemistry / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D017354:Point Mutation; D017386:Sequence Homology, Amino Acid; D019098:Telomerase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*
D017931:DNA Primers; D004273:DNA, Neoplasm; D004268:DNA-Binding Proteins; D054303:HIV Reverse Transcriptase; C509186:TERT protein, human; D019098:Telomerase