<p>Bacterial resistance to antibiotics has become a serious problem. This is due to the remarkable ability of bacteria to respond and rapidly adapt to environmental changes. Integrons are elements with the capacity for gene capture by an integron-encoded site-specific recombinase called IntI. IntI binds and acts at the recombination sites, <i>attI </i>and<i> attC</i> resulting in excision and integration of short DNA elements called gene cassettes carrying an <i>attC</i> site in the 3’ end. Several families of antibiotic resistance genes are borne on gene cassettes in integrons connected to mobile elements. Other cassettes reside in the larger and ancestral superintegrons located on chromosomes in both pathogenic and environmental bacteria. Due to their close connection with lateral gene transfer systems, it is possible that integrons are functionally dependent on those networks. This work presents arguments for such connections. The<i> attC</i> of the <i>aadA1-qacE</i> cassette junction in Tn<i>21</i> was characterized in detail. Like other <i>attC</i> sites, it contains two pairs of inverted repeats and is almost palindromic. By using electrophoretic mobility shift assays, this study showed that IntI1 binds only to the bottom strand of <i>attC</i>. Upon folding the strand into a hairpin, a few chiral hairpin distortions define both the strand choice and also the appropriate orientation of the highly symmetrical site. Structural recognition also explains the wide sequence variation among <i>attC</i> sites. We have documented the initial cleavage step in recombination in IntI extracts and integrase levels in extracts were evaluated by a new method. Mutagenesis and homology modelling were performed to find amino acid residues in IntI1 that are important for recognition of <i>attC</i> hairpin-DNA. Comparisons were made with other tyrosine family members to explain how integron integrases differ in site-recognition and also in their mechanism of strand exchange.</p>
| Identifer | oai:union.ndltd.org:UPSALLA/oai:DiVA.org:uu-7429 |
| Date | January 2007 |
| Creators | Johansson, Carolina |
| Publisher | Uppsala University, Department of Medical Biochemistry and Microbiology, Uppsala : Acta Universitatis Upsaliensis |
| Source Sets | DiVA Archive at Upsalla University |
| Language | English |
| Detected Language | English |
| Type | Doctoral thesis, comprehensive summary, text |
| Relation | Digital Comprehensive Summaries of Uppsala Dissertations from the Faculty of Medicine, 1651-6206 ; 214 |
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