\documentclass{beamer}

\usetheme{Pittsburgh}
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\title{Kravats et al.'s ``Unfolding and translocation pathway of
  substrate protein controlled by structure in repetitive allosteric
  cycles of the ClpY ATPase''}
\author{Trevor King}
\institute{Drexel University} \date{\today}

\begin{document}

\begin{frame}
  \titlepage
\end{frame}

\begin{frame}
  \frametitle{Clp macromachines}
  \begin{itemize}
    \item Clp: caseinolytic protease
    \item Found in all forms of life
    \item Involved with selective destruction of proteins
    \item Clp ATPases force protein unfolding and translocation
      through narrow pores
    \item Functional ClpATPases are homohexameric assemblies (form in
      presence of ATP)
  \end{itemize}
\end{frame}

\begin{frame}
  \frametitle{The Clp ATPase channel}
  \begin{itemize}
    \item Each monomer has one or two AAA domains for binding nucleotides
    \item Core of hexamer forms a channel with a minimum diameter of
      $9$-$15\AA$
    \item Each AAA domain provides a diaphram-forming central loop in
      the channel
    \item Loops have a conserved aromatic-hydrophobic dipeptide
      flanked by glycines
    \item Loops involved with regulated substrate selection, binding
      to tags fused to target proteins
    \item Protein unfolded to pass through channel, often involving
      ATP hydrolysis
    \item Mechanical force comes from nucleotide-dependent loop
      displacement
  \end{itemize}
\end{frame}

\begin{frame}
  \frametitle{Possible mechanisms}
  Paddles cannot generate enough force to match AFM-measured unfolding forces.

  What's going on?
  \begin{itemize}
    \item Loop padling action
    \item Distinct unfolding mechanisms (?) from AFM-based studies
    \item Constant force + periodic modulation to shift AFM unfolding pathway
  \end{itemize}
\end{frame}

\begin{frame}
  \frametitle{This paper}
  Approach:
  \begin{itemize}
    \item Coarse-grained simulation
    \item Check each mechanism to estimate timescale
  \end{itemize}

  Results:
  \begin{itemize}
    \item Obligatory unfolding intermediate (C-terminal unraveling)
    \item Non-native substrate conformations stabilized by ClyY at
      pore entrance
    \item Weaker loop forces sufficient to unfold and translocate
      stabilized substrate
  \end{itemize}
\end{frame}

\begin{frame}
  \frametitle{Snapshots of unfolding and translocation}
  \begin{center}
    \includegraphics[height=0.9\textheight]{figure/fig-1}
  \end{center}
\end{frame}

\begin{frame}
  \frametitle{Simulating AFM unfolding}
  \begin{itemize}
    \item Fix N terminus, pull C terminus at constant velocity
    \item Identify major pathway (75\%) where N-terminal helix
      unravels first, followed by C-terminal helix and intermediate
      helicies.
    \item Simulated peak forces $\sim 50$-$100$ pN.  (experiment:
      $25$-$35$ pN)
    \item Similar pathway to implicit-solvent simulations
  \end{itemize}
\end{frame}

\begin{frame}
  \frametitle{Is ATP required?}
  \begin{itemize}
    \item Presented HBP to nonallosteric, open-state ClpY ring
    \item SsrA tag binds to central channel loops
    \item No HBP translocation, HPB retains native structure
  \end{itemize}
\end{frame}

\begin{frame}
  \frametitle{ATP-driven pumping}
  \begin{itemize}
    \item Simulate ATP pumping by forcing subunit motion (more later)
    \item Generates periodic force driving substrate
    \item Brings folded HBP up against channel loops
    \item Starts unraveling C terminus
  \end{itemize}
\end{frame}

\begin{frame}
  \frametitle{Fraction of native contacts}
  Blue: after binding, black: after 50 ATP-driven cycles, red: after
  50 cycles with ClpQ, green: native
  \begin{center}
    \includegraphics[width=0.8\textwidth]{figure/fig-2}
  \end{center}
\end{frame}

\begin{frame}
  \frametitle{Translocation and radius of gyration}
  \begin{center}
    (A) Radius of HBP under ATP-driven ClpY;
    (B) Length of HBP translocated in the pore
    \includegraphics[width=0.9\textwidth]{figure/fig-3}
  \end{center}
\end{frame}

\begin{frame}
  \frametitle{Fraction native contacts and radius of gyration}
  \begin{center}
    \includegraphics[width=0.9\textwidth]{figure/fig-4}
  \end{center}
\end{frame}

\begin{frame}
  \frametitle{}
  \begin{center}
    \includegraphics[width=0.9\textwidth]{figure/fig-5}
  \end{center}
\end{frame}

\end{document}
