| |
| author |
Zarina Akbary
| | title |
Ketoreductase Activity of 45-million-year-old Yeast Strains Revived from Amber
| | abstract |
Industrially relevant enzymes, such as ketoreductases, are often identified by searching
through bioinformatic databases. Those enzyme databases are limited by our lack of
knowledge but can be expanded upon through our exploration of biodiversity. Notably,
biodiversity is not limited to contemporary organisms spatially distributed across the globe,
but can also encompass the ancestors of existing ones. Thus, the investigation of temporal
diversity may be as important an avenue for enzymatic discovery as the exploration of
exotic or under-studied microbial communities. In this work, we screened four 45-million-
year-old yeast strains previously revived from fossilized amber for ketoreductase activity
using a panel of representative ketone substrates. After successful demonstration of native
whole-cell ketoreductase activity, we annotated a previously sequenced genome of one of
those ancient strains, SC108, and identified eight aldo-ketoreductases (AKR) of interest.
We successfully cloned five of these AKRs from SC108 into an expression vector to
generate affinity-tagged fusion proteins. Screening the clones expressing each of the AKRs
demonstrated that these five enzymes contributed, in part, to the ketoreductase activity seen
with the native whole cells. The chirality of the alcohol products generated by the AKRs
were determined using an HPLC-based method and suggested three of these ancient
enzymes exhibited anti-Prelog enantioselectivity, and one enzyme exhibited Prelog
enantioselectivity. The enantioselectivity of one of the enzymes could not be determined.
| | school |
The College of Liberal Arts, Drew University
| | degree |
B.A. (2019)
|
| advisor |
Neal Connors
|
| committee |
Joanna Miller
Kimberly Choquette
|
| full text | ZAkbary.pdf - requires Drew uLogin |
| |
