#chemistry

ClpXP degradation system in E. coli; a study of its energy sources and its applications in managing the expression levels of targeted membrane and soluble proteins

Abstract:
 

 ClpXP is an Escherichia coli protease that carryout energy-dependent intracellular proteolysis. In recent years, this system has been widely studied due to its importance as a protein regulatory machinery and a virulence factor.  Protein substrates of ClpXP contain degrons with a specific protein sequence. SsrA tag is one of the five degrons known to subject proteins for ClpXP degradation. SsrA is an 11 amino acid peptide added to the C-terminus of nascent polypeptide chains translated from aberrant messenger RNAs lacking stop codons via a process called trans-translation.

ClpXP was known to targets only cytosolic proteins with degrons until recently, AcrB, an E. coli membrane protein was found to be degraded by ClpXP when it is tagged by ssrA peptide, which leads to the speculation that ClpXP is capable of degrading membrane proteins.   However, this speculation was challenged with the finding that ssrA tagging of ProW1−182, a different inner-membrane protein resulted in degradation by AAA+ membrane protease FtSH. We report that the membrane substrates of ClpXP bear long c-terminal cytoplasmic domains while metastable proteins lacking cytoplasmic domains are degraded by FtsH. For instance, ssrA tagged Aquaporin-Z (AqpZ), a stable tetrameric membrane protein lacking a c-terminal cytoplasmic domain is subjected to degradation by neither ClpXP nor FtsH. Nevertheless, when the c-terminus of AqpZ is fused with ssrA tagged Cyan fluorescent protein ClpXP degrades the resulted fusion protein while truncated metastable version, AqpZ 1-155 is degraded by FtSH.

This presentation also emphasizes our attempt to unravel the possible effect of proton motive force on the activity of ClpXP. We used Carbonyl cyanide m-chlorophenyl hydrazone (CCCP) to disrupt the proton motive force. Our results suggest that degradation of soluble protein substrates such as GFP-ssrA, MurA-ssrA, Chloramphenicol acetyltransferase ssrA are not affected by CCCP. However, degradation of membrane protein substrates by ClpXP is diminished in the presence of CCCP. We speculate that either the proton motive force or ATP provided from oxidative phosphorylation is essential, or both are important for ClpXP to degrade membrane proteins. 

It has been shown that the TolC is not a good target for inhibition of multidrug efflux of antibiotic-resistant bacteria as the bacterial susceptibility to antibiotics was not affected even when a significant amount of TolC is depleted.  TolC is a membrane protein channel that functions in conjunction with transporters and membrane fusion proteins and provides a pathway to expel antibiotics across the E. coli outer membrane.  AcrAB-TolC multidrug efflux pump is one such example where TolC cooperates with AcrB transporter and AcrA membrane fusion protein.   We report that the depletion of the number of copies of AcrB makes bacteria highly susceptible to antibiotics. We utilized ClpXP degradation system to regulate the copy number of AcrB in the cell. Our results show that AcrB is an excellent target for inhibiting multidrug efflux, and ClpXP is an excellent tool to regulate antibiotic target proteins for research purposes.
Date: 
Friday, April 23, 2021 - 9:00am to 10:00am
Location: 
Zoom
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2021 Regional Undergraduate Poster Competition

Find details of the event and registration here.

To view a copy of the 2019 abstract booklet, click here.

Note to UK students: Students in CHE 395 planning to graduate or otherwise conclude their research are required to participate in the Poster Session if they have not done so in the past. 

Schedule of Events:
10:00am - Zoom Check-In and Set Up
10:30 - 12:00pm - Group A Presents
1:00pm - 2:30pm - Group B Presents
3:30pm - Awards Presented

First Prize

$300

Second Prize

$200

Honorable Mention

3 @ $100

Recent winners include students from:

Belmont University
Berea College
Centre College
Indiana State University
Indiana University
Indiana University Kokomo
Marshall University
Rose-Hulman Institute of Technology
Transylvania University
University of Kentucky
Western Kentucky University
 

We thank the Lexington Section of the American Chemical Society for graciously funding the awards for this poster session.

Please contact the department if you have questions.

Date: 
Friday, April 23, 2021 - 10:00am to 4:00pm
Location: 
Virtual
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A THEORETICAL AND EXPERIMENTAL STUDY OF CHARGE TRANSPORT IN ORGANIC THERMOELECTRIC MATERIALS AND CHARGE TRANSFER STATES IN ORGANIC PHOTOVOLTAICS

Abstract: Application of organic electronics have increased significantly over the past two decades. Organic materials can be used in flexible devices with cheaper cost of fabrication, yet in most cases the devices suffer from poor performance and stability. Investigating doping mechanism, charge transport and charge transfer in such materials can help us to understand the origin of these issues and later resolve them. In this dissertation, organic materials are used in three different device structures to investigate charge transport and charge transfer. Chemically doped pi-conjugated polymers are promising materials to be used in thermoelectric (TE) devices, yet their application is limited by their low performance. Blending two polymers is a simple way to change the properties of the TE devices. Here we used a simple analytical model to calculate TE properties of polymer blend by taking into account for energetic disorder, energetic offset between two polymers and localization length which proposed TE performance of polymer blend can exceed the individual ones at specific blends of two polymers. We showed these improvements are achievable by experimentally testing TE properties of selected polymer blends. Further, to investigate the doping mechanism in polymers, we used organic electrochemical transistors to investigate the effect of anion size on polaron delocalization and the thermoelectric properties of single polymers. This device structure allowed us to control the charge carrier concentration with minimizing the effects on the film morphology.

In organic photovoltaics (OPVs), upon fluorination of donor molecules the performance of device increases in most cases. So, we investigated the charge transfer state energy between the electron donor anthradithiophene (ADT) and the electron acceptor C60 upon halogenation of the ADT molecule. Interfacial energetics and charge transfer state energies between donor and acceptor are crucial to performance of these devices. We probe interfacial energetics of donor/acceptor interfaces with Ultraviolet photoelectron spectroscopy (UPS) charge transfer state energies with sensitive External Quantum Efficiency (EQE) setup both in bilayer and bulk heterojunction device structure. These measurements coupled with DFT calculations allowed us to explain the effects of halogenation on the OPV devices characteristics. Investigating charge transfer states energies, charge transport and doping mechanism in organic materials allow us to improve the performance of organic based electronics and also propose new applications for these family of materials.

 

 

Date: 
Friday, October 23, 2020 - 9:00am to 10:00am
Location: 
Zoom
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Dr. Sean Parkin Named Section Editor of Acta Crystallographica E

Congratulations to Dr. Sean Parkin, named

Section Editor of Acta Crystallographica E!

Read the full story here.

The Story of a 1949 UK Chemistry Graduate

It was 1949, World War II had ended and twice as many students were enrolled in universities across the country compared to pre-war enrollment, many were on the GI Bill. I was one of those June 1949 GI Bill seniors, graduating from UK with a BS degree in physical chemistry. My name is Alan Veith.

Catalent Info Session

Date: 
Thursday, October 24, 2019 - 3:45pm to 5:45pm
Location: 
Whitehall Classroom Building, Room 331
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Photocatalytic reductions occur in slow time scales

Finding the time scale for the effective transfer of electrons is not an easy task.

Infectious Diseases, Auto-Immune Diseases, and Opportunities for Biophysical Chemistry

We present examples from our group where biophysical chemistry impacts unsolved problems in infectious diseases and auto-immune diseases. We start with bacterial biofilms, which are structured multi-cellular communities that are fundamental to the biology and ecology of bacteria. By using population tracking algorithms, we dissect bacterial social behavior at the single cell level.  We will also discuss how we can learn from innate immunity peptides to renovate antibiotic design, and make precision antibiotics and antibiotics against persister bacterial populations. Finally, we examine the pathological role of antimicrobial peptides in a range of autoimmune disorders.

 

Date: 
Friday, April 8, 2016 - 4:00pm to 5:00pm
Location: 
CP - 114
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