Identification of a new variant CYP2D6 allele with a single base deletion in exon 3 and its association with the poor metabolizer phenotype

AbstractDbh is a Y-family translesion DNA polymerase from Sulfolobus acidocaldarius, an archaeal species that grows in harsh environmental conditions. Biochemically, Dbh displays a distinctive mutational profile, creating single-base deletion mutations at extraordinarily high frequencies (up to 50%) in specific repeat sequences. In cells, however, Dbh does not appear to contribute significantly to spontaneous frameshifts in these same sequence contexts. This suggests that either the error-prone DNA synthesis activity of Dbh is reduced in vivo and/or Dbh is restricted from replicating these sequences. Here, we test the hypothesis that the propensity for Dbh to make single base deletion mutations is reduced through interaction with the S. acidocaldarius heterotrimeric sliding clamp processivity factor, PCNA-123. We first confirm that Dbh physically interacts with PCNA-123, with the interaction requiring both the PCNA-1 subunit and the C-terminal 10 amino acids of Dbh, which contain a predicted PCNA-interaction peptide (PIP) motif. This interaction stimulates the polymerase activity of Dbh, even on short, linear primer-template DNA by increasing the rate of nucleotide incorporation. This stimulation requires an intact PCNA-123 heterotrimer and a DNA duplex length of at least 18 basepairs, the minimal length predicted from structural data to bind to both the polymerase and the clamp. Finally, we find that PCNA-123 increases the fidelity of Dbh on a single-base deletion hotspot sequence 3-fold by promoting an increase in the rate of correct, but not incorrect, nucleotide addition and propose that PCNA-123 induces Dbh to adopt a more active conformation that is less prone to creating deletions during DNA synthesis.HighlightsPCNA increases the fidelity of Dbh polymerase on a deletion-hotspot sequence.The interaction stimulates incorporation of the correct, but not incorrect, nucleotide.A minimal duplex length of 18 bp is required for PCNA to stimulate polymerase activity.Structural modeling suggests that PCNA induces a conformational change in Dbh.

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Metoprolol α-Hydroxylation Polymorphism in the San Bushmen of Southern Africa

Human Toxicology ◽

10.1177/096032718900800107 ◽

1989 ◽

Vol 8 (1) ◽

pp. 39-43 ◽

Cited By ~ 12

Author(s):

De K. Sommers ◽

J. Moncrieff ◽

J. Avenant

Keyword(s):

Metabolic Rate ◽

Frequency Distribution ◽

Southern Africa ◽

Bimodal Distribution ◽

Oxidative Status ◽

Urine Collection ◽

Rate Data ◽

Poor Metabolizer ◽

Unimodal Distribution ◽

The Poor

1 The metabolic oxidation of metoprolol has been studied in a group of 98 San Bushmen. 2 The amounts of metoprolol and α-hydroxy metoprolol excreted in 0-8 h urine collection, after dosing with 100 mg metoprolol, were measured and the metabolic ratio (% dose excreted as metoprolol/% dose excreted as α-hydroxy metoprolol) calculated. 3 Frequency distribution and probit plots of the metabolic rate data showed a bimodal distribution with 4.1% of the population exhibiting slow metabolism with an MR > 10. 4 These results are much less than found in Caucasians (8.4%) but very different from the unimodal distribution found for Nigerians. 5 A previous study in the same group of Bushmen had revealed that 18 of 96 subjects were poor or non-metabolizers of debrisoquine to 4-hydroxy debrisoquine, but only one of the poor metoprolol metabolizers was a poor metabolizer of debrisoquine. 6 On the basis of these results, the claim of debrisoquine type of polymorphism for β-adrenoceptor antagonists found in Caucasians cannot be extrapolated to the San Bushmen, and one must query the use of debrisoquine as measure of oxidative status in any group other than Caucasians.

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