RESUMO
Functioning DNA repair capabilities are vital for organisms to ensure that the biological information is preserved and correctly propagated. Disruptions in DNA repair pathways can result in the accumulation of DNA mutations, which may lead to onset of complex disease such as cancer. The discovery and characterization of cancer-related biomarkers may allow early diagnosis and targeted treatment, which could significantly contribute to the survival rates of cancer patients. To this end, we have applied a hypothesis driven bioinformatics approach to identify biomarkers related to 25 different DNA repair enzymes, in combination with structural analysis of six selected missense mutations of newly discovered SNPs that are associated with cancer phenotypes. Our search on 8 distinct cancer databases uncovered 43 missense SNPs that statistically significantly associated at least one phenotype. Moreover, nine of these missense SNPs are statistically significantly associated with two or more cancers. In addition, we have performed classical molecular dynamics to characterize the impact of rs10018786 on POLN, which results in the M310 L Pol ν variant, and rs3218784 on POLI, which results in the I236 M Pol ι. Our results suggest that both of these cancer-associated variants result in noticeable structural and dynamical changes compared with their respective wild-type proteins.
Assuntos
Biomarcadores Tumorais/genética , Reparo do DNA/genética , Bases de Dados Genéticas , Fenótipo , DNA Polimerase I/química , DNA Polimerase I/genética , DNA Polimerase I/metabolismo , Simulação de Dinâmica Molecular , Mutação , Mutação de Sentido Incorreto , Polimorfismo de Nucleotídeo Único , Conformação ProteicaRESUMO
Structural integrity of DNA molecules is necessary for their information storage function. Cells rely on a number of pathways to ensure that the damage to DNA induced by endogenous and exogenous reagents is repaired. AlkD, a base excision enzyme, removes a damaged nucleobase by cleaving a glycosidic bond. Unlike many other base excision enzymes, AlkD does not flip a damaged nucleobase into a designated reaction pocket, and as such can repair nucleobases with larger adducts, such as yatakemycin. In this study, the structure and dynamics of AlkD have been investigated by classical molecular dynamics simulations. Several systems including apo-AlkD, and AlkD in complex with DNA, both with and without the yatakemycin adduct have been simulated. Comparison of the results for the apo-AlkD with AlkD with substrate (damaged or undamaged) indicates a high degree of motion of helix αB in apo-AlkD, whereas this helix is observed to form various contacts when the substrate is bound. The calculated results are consistent with previous experimental studies that have suggested various residues involved in damage recognition, DNA binding, and base excision catalysis.
RESUMO
The search for prostate cancer biomarkers has received increased attention and several DNA repair related enzymes have been linked to this dysfunction. Here we report a targeted search for single nucleotide polymorphisms (SNPs) and functional impact characterization of human ALKBH family dioxygenases related to prostate cancer. Our results uncovered a SNP of ALKBH7, rs7540, which is associated with prostate cancer disease in a statistically significantly manner in two separate cohorts, and maintained in African American men. Comparisons of molecular dynamics (MD) simulations on the wild-type and variant protein structures indicate that the resulting alteration in the enzyme induces a significant structural change that reduces ALKBH7's ability to bind its cosubstrate. Experimental spectroscopy studies with purified proteins validate our MD predictions and corroborate the conclusion that this cancer-associated mutation affects productive cosubstrate binding in ALKBH7.
Assuntos
Enzimas AlkB/genética , Ácidos Cetoglutáricos/química , Proteínas Mitocondriais/genética , Polimorfismo de Nucleotídeo Único/genética , Neoplasias da Próstata/etnologia , Neoplasias da Próstata/genética , Negro ou Afro-Americano/estatística & dados numéricos , Sítios de Ligação , Biomarcadores Tumorais/química , Biomarcadores Tumorais/genética , Ativação Enzimática , Marcadores Genéticos/genética , Predisposição Genética para Doença/etnologia , Predisposição Genética para Doença/genética , Humanos , Masculino , Simulação de Dinâmica Molecular , Oxigênio/química , Prevalência , Ligação Proteica , Fatores de Risco , Especificidade por Substrato , Estados Unidos/epidemiologia , Estados Unidos/etnologiaRESUMO
The ability to repair DNA is important for the conservation of genetic information of living organisms. Cells have a number of ways to restore damaged DNA, such as direct DNA repair, base excision repair, and nucleotide excision repair. One of the proteins that can perform direct repair of DNA bases is Escherichia coli AlkB. In humans, there are 9 identified AlkB homologs, including AlkB homolog 1 (ALKBH1). Many of these proteins catalyze the direct oxidative dealkylation of DNA and RNA bases and, as such, have an important role in repairing DNA from damage induced by alkylating agents. In addition to the dealkylase activity, ALKBH1 can also function as an apyrimidinic/apurinic lyase and was proposed to have a distinct lyase active site. To our knowledge, no crystal structure or complete homology model of ALKBH1 protein is available. In this study, we have used homology modeling to predict the structure of ALKBH1 based on AlkB and Duffy-binding-like domain crystal structures as templates. Molecular dynamics simulations were subsequently performed on the predicted structure of ALKBH1. The positions of two disulfide bonds or a zinc-finger motif and a disulfide bond were predicted and the importance of these features was tested by mutagenesis. Possible locations for the lyase active site are proposed based on the analysis of our predicted structures and previous experimental results.
