The N-terminus of Himar1 mariner transposase mediates multiple activities during transposition.
Mariner family transposons are perhaps the most widespread transposable elements of eukaryotes. While we are beginning to understand the precise mechanism of transposition of these elements, the structure of their transposases are still poorly understood. We undertook an extensive mutagenesis of the N-terminal third of the transposase of the Himar1 mariner transposon to begin the process of determining the structure and evolution of mariner transposases. N and C-terminal deletion analyses localized the DNA binding domain of Himar1 transposase to the first 115 amino acids. Alanine scanning of 23 selected sites within this region uncovered mutations that not only affected DNA binding but DNA cleavage as well. The behavior of other mutations strongly suggested that the N-terminus is also involved in multimerization of the transposase on a single inverted terminal repeat and in paired ends complex formation which brings together the two ends of the transposon. Finally, two hyperactive mutations at conserved sites suggest that mariner transposases are under a pattern of stabilizing selection in nature with regard to how efficiently they mediate transposition, resulting in a population of "average" transposons.
Butler
Matthew G
MG
Department of Human Genetics, University of Michigan, Ann Arbor, MI 48109, USA.
Chakraborty
Sangita A
SA
Lampe
David J
DJ
D000409:Alanine / Q000032:analysis; D000595:Amino Acid Sequence; D000818:Animals; D017124:Conserved Sequence; D004251:DNA Transposable Elements / Q000502:physiology*; D004268:DNA-Binding Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism* / Q000502:physiology*; D003850:Deoxyribonuclease I / Q000378:metabolism; D024202:Electrophoretic Mobility Shift Assay; D004926:Escherichia coli; D014644:Genetic Variation; D008954:Models, Biological; D008969:Molecular Sequence Data; D009117:Muscidae / Q000235:genetics / Q000378:metabolism / Q000382:microbiology; D050505:Mutant Proteins / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary / Q000502:physiology; D017384:Sequence Deletion; D017386:Sequence Homology, Amino Acid; D013045:Species Specificity; D019895:Transposases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism* / Q000502:physiology*
D004251:DNA Transposable Elements; D004268:DNA-Binding Proteins; D050505:Mutant Proteins; C111652:mariner transposases; D019895:Transposases; D003850:Deoxyribonuclease I; D000409:Alanine