Molecular mechanisms of template-independent CCA-addition (14218)
Every tRNA has the CCA sequence at its 3'-terminus (CCA-3' at positions 74 - 76; C74C75A76-3'). The CCA-3' moiety is required for amino acid attachment (aminoacylation) onto the 3'-end of the tRNA by aminoacyl-tRNA synthetases, and for peptide-bond formation on the ribosome. The universal CCA-3' of tRNA is built and/or synthesized by the CCA-adding enzyme, CTP:(ATP) tRNA nucleotidyltransferase. This RNA polymerase has no nucleic acid template, but faithfully synthesizes the defined CCA sequence on the 3'-terminus of tRNA at one time, using CTP and ATP as substrates. The mystery of CCA-addition without a nucleic acid template by unique RNA polymerases has long fascinated researchers in the field of RNA enzymology.
The CCA-adding enzymes are classified into two classes (class-I and class-II), based on the sequence alignments. The archaeal CCA-adding enzymes belong to class-I, and share sequence similarity with the eukaryotic PAPs. On the other hand, the eubacterial/eukaryotic CCA-adding enzymes belong to class-II, and share sequence similarity with the eubacterial PAPs. Although both enzyme classes catalyze the same reaction in defined fashions, significant amino acid similarities are not readily apparent between the two classes of CCA-adding enzymes. Only local similarities around the active site signatures have been identified. In most organisms, the CCA-3' is synthesized by a single enzyme that can add the CCA-3' at one time. However, in some eubacteria, the CCA-3' is synthesized by two distinct, but closely related, class-II enzymes. One adds CC and the other adds A, and the CCA-3' is synthesized by these two enzymes in a collaborative manner.
Over the past twelve years, the crystal structures of class-I and class-II enzymes and their complexes with nucleotide(s) and/or various tRNAs or mini-helices have been reported. The structural information has solved most of the long-standing mysteries, but not all yet, about the mechanism of the remarkable, template-independent CCA-adding enzyme and its relatives. In this talk, we present the current understanding of the mechanism of CCA-addition by the CCA-adding enzyme, and its related enzymes, based on their crystal structures.
The CCA-adding enzymes are classified into two classes (class-I and class-II), based on the sequence alignments. The archaeal CCA-adding enzymes belong to class-I, and share sequence similarity with the eukaryotic PAPs. On the other hand, the eubacterial/eukaryotic CCA-adding enzymes belong to class-II, and share sequence similarity with the eubacterial PAPs. Although both enzyme classes catalyze the same reaction in defined fashions, significant amino acid similarities are not readily apparent between the two classes of CCA-adding enzymes. Only local similarities around the active site signatures have been identified. In most organisms, the CCA-3' is synthesized by a single enzyme that can add the CCA-3' at one time. However, in some eubacteria, the CCA-3' is synthesized by two distinct, but closely related, class-II enzymes. One adds CC and the other adds A, and the CCA-3' is synthesized by these two enzymes in a collaborative manner.
Over the past twelve years, the crystal structures of class-I and class-II enzymes and their complexes with nucleotide(s) and/or various tRNAs or mini-helices have been reported. The structural information has solved most of the long-standing mysteries, but not all yet, about the mechanism of the remarkable, template-independent CCA-adding enzyme and its relatives. In this talk, we present the current understanding of the mechanism of CCA-addition by the CCA-adding enzyme, and its related enzymes, based on their crystal structures.