Nadhan, Navien Tholasi
(2022)
In Silico Selection And Validation Of
Dna Aptamer Against Progesterone
Receptor Dna Binding Domain.
Masters thesis, Universiti Sains Malaysia.
Abstract
Progesterone receptor plays an important role in the progression of breast
cancer. Currently, antibody-based Immunohistochemistry is used in pathological
assessment of PR levels for the detection of breast cancer. The shortcomings
associated with antibodies pave the path to use aptamers as the alternatives.
Aptamers are single-stranded DNA or RNA oligonucleotides generated by SELEX
that are capable of binding to their cognate target molecules with high affinity and
specificity based on their unique structural folding capacity. The tediousness and
rigor associated with certain steps of the conventional SELEX intensify the efforts to
select DNA aptamers using in silico-docking approach. That said, we report an in
silico selection and validation of DNA aptamer to the progesterone receptor DNA
binding domain (PR DBD) using ssDNA sequences derived from human
progesterone response elements (PREs). Firstly, a library of sixty-four different nearnative
ssDNA analogs of the corresponding PRE sequences was designed and
subjected to secondary and tertiary structural determination. After that, docking
between the ssDNA tertiary structures with the PR DBD was carried out using
PatchDock. The sequence with the highest docking score was chosen as the aptamer
candidate and further validated by in vitro direct ELASA. Among the candidates, we
selected the ssDNA sequence (PRDBDapt17; 5′- AGAACAGCGTGTTCT -3′),
which showed the highest docking scores of 11334 as a promising PR DBD binding
aptamer. In addition, the PRDBDapt17 detected recombinant PR DBD in direct
ELASA with a limit of detection of 3.91 nM.
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