Inner mitochondrial membrane resident 3-beta hydroxysteroid dehydrogenase2 (3βHSD2) requires chaperone-assisted folding and release for steroidogenic activity

Program: Abstracts - Orals, Featured Poster Presentations, and Posters
Session: MON 515-547-Female Reproductive Endocrinology
Basic/Translational
Monday, June 17, 2013: 1:45 PM-3:45 PM
Expo Halls ABC (Moscone Center)

Poster Board MON-520
Maheshinie Rajapaksha*1, Manoj Prasad2, James L Thomas3, Randy Whittal4 and Himangshu S Bose5
1Mercer University School of Medicine, Savannah, GA, 2Mercer University, Savannah, GA, 3Mercer Univ Sch of Med, Macon, GA, 4University of Alberta, Edmonton, Canada, 5Mercer Univ Schl of Med, Savannah, GA
The inner mitochondrial membrane resident 3-beta hydroxysteroid dehydrogenase2 (3bHSD2) synthesizes progesterone and androstenedione through dehydrogenase and isomerase activities, necessitating the protein to undergo a reversible conformational change. We hypothesized that chaperones assist 3bHSD2 to switch between the conformations for dual functionality. Circular dichroism showed that addition of LM (Lauryl Maltoside) increased ellipticity at both the p-p* and n-p* positions, which over-time reverted to control levels, suggesting the formation of a stable but reversible conformation possibly due to the hydrophobic interaction of the protein with detergent micelles. Using ANS (8-anilino-1-naphthalenesulfonic acid), we found a LM-mediated decrease in apparent binding, from 2.36x106 to 5.40x104 M-1, as ANS molecules may no longer have access to the hydrophobic regions of 3bHSD2. In presence of chaperones metabolic conversion was increased more with NP40 than LM and then remained unchanged up to the miceller limit possibly because hydrophobic regions of the protein interacted with detergent micelles.  This observation supports the idea that detergents act as molecular chaperones to assist 3bHSD2 in forming stable complexes, which in turn promotes proper folding and function. Finger printing illustrated that LM incubation resulted in four bands of molecular mass from 39-13 kDa.  Space filling modeling demonstrates that association with chaperones likely exposed the hydrophobic region, leading to its proteolysis.  We conclude that chaperones help 3bHSD2 to refold in order to rejuvenate, contributing to the ability of cells to rapidly produce steroids when needed.

Nothing to Disclose: MR, MP, JLT, RW, HSB

*Please take note of The Endocrine Society's News Embargo Policy at http://www.endo-society.org/endo2013/media.cfm

Sources of Research Support: NIH