Paul Blount

Publications (67) View all

• Article: Chimeras Reveal a Single Lipid-Interface Residue that Controls MscL Channel Kinetics as well as Mechanosensitivity.

  Li-Min Yang, Dalian Zhong, Paul Blount
  [show abstract] [hide abstract]
  ABSTRACT: MscL, the highly conserved bacterial mechanosensitive channel of large conductance, serves as an osmotic "emergency release valve," is among the best-studied mechanosensors, and is a paradigm of how a channel senses and responds to membrane tension. Although all homologs tested thus far encode channel activity, many show functional differences. We tested Escherichia coli and Staphylococcus aureus chimeras and found that the periplasmic region of the protein, particularly E. coli I49 and the equivalent S. aureus F47 at the periplasmic lipid-aqueous interface of the first transmembrane domain, drastically influences both the open dwell time and the threshold of channel opening. One mutant shows a severe hysteresis, confirming the importance of this residue in determining the energy barriers for channel gating. We propose that this site acts similarly to a spring for a clasp knife, adjusting the resistance for obtaining and stabilizing an open or closed channel structure.
  Cell reports. 02/2013; 3(2):520-7.
• Article: Structure and molecular mechanism of an anion-selective mechanosensitive channel of small conductance.

  Xiaozhe Zhang, Jingjing Wang, Yue Feng, Jingpeng Ge, Wenfei Li, Wending Sun, Irene Iscla, Jie Yu, Paul Blount, Yang Li, Maojun Yang
  [show abstract] [hide abstract]
  ABSTRACT: Mechanosensitive (MS) channels are universal cellular membrane pores. Bacterial MS channels, as typified by MS channel of small conductance (MscS) from Escherichia coli (EcMscS), release osmolytes under hypoosmotic conditions. MS channels are known to be ion selective to different extents, but the underlying mechanism remains poorly understood. Here we identify an anion-selective MscS channel from Thermoanaerobacter tengcongensis (TtMscS). The structure of TtMscS closely resembles that of EcMscS, but it lacks the large cytoplasmic equatorial portals found in EcMscS. In contrast, the cytoplasmic pore formed by the C-terminal β-barrel of TtMscS is larger than that of EcMscS and has a strikingly different pattern of electrostatic surface potential. Swapping the β-barrel region between TtMscS and EcMscS partially switches the ion selectivity. Our study defines the role of the β-barrel in the ion selection of an anion-selective MscS channel and provides a structural basis for understanding the ion selectivity of MscS channels.
  Proceedings of the National Academy of Sciences 10/2012; · 9.68 Impact Factor
• Article: Sensing and responding to membrane tension: the bacterial MscL channel as a model system.

  Irene Iscla, Paul Blount
  [show abstract] [hide abstract]
  ABSTRACT: Mechanosensors are important for many life functions, including the senses of touch, balance, and proprioception; cardiovascular regulation; kidney function; and osmoregulation. Many channels from an assortment of families are now candidates for eukaryotic mechanosensors and proprioception, as well as cardiovascular regulation, kidney function, and osmoregulation. Bacteria also possess two families of mechanosensitive channels, termed MscL and MscS, that function as osmotic emergency release valves. Of the two channels, MscL is the most conserved, most streamlined in structure, and largest in conductance at 3.6 nS with a pore diameter in excess of 30 Å; hence, the structural changes required for gating are exaggerated and perhaps more easily defined. Because of these properties, as well as its tractable nature, MscL represents a excellent model for studying how a channel can sense and respond to biophysical changes of a lipid bilayer. Many of the properties of the MscL channel, such as the sensitivity to amphipaths, a helix that runs along the membrane surface and is connected to the pore via a glycine, a twisting and turning of the transmembrane domains upon gating, and the dynamic changes in membrane interactions, may be common to other candidate mechanosensors. Here we review many of these properties and discuss their structural and functional implications.
  Biophysical Journal 07/2012; 103(2):169-74. · 3.65 Impact Factor
• Article: The dynamics of protein-protein interactions between domains of MscL at the cytoplasmic-lipid interface.

  Irene Iscla, Robin Wray, Paul Blount
  [show abstract] [hide abstract]
  ABSTRACT: The bacterial mechanosensitive channel of large conductance, MscL, is one of the best characterized mechanosensitive channels serving as a paradigm for how proteins can sense and transduce mechanical forces. The physiological role of MscL is that of an emergency release valve that opens a large pore upon a sudden drop in the osmolarity of the environment. A crystal structure of a closed state of MscL shows it as a homopentamer, with each subunit consisting of two transmembrane domains (TM). There is consensus that the TM helices move in an iris like manner tilting in the plane of the membrane while gating. An N-terminal amphipathic helix that lies along the cytoplasmic membrane (S1), and the portion of TM2 near the cytoplasmic interface (TM2ci), are relatively close in the crystal structure, yet predicted to be dynamic upon gating. Here we determine how these two regions interact in the channel complex, and study how these interactions change as the channel opens. We have screened 143 double-cysteine mutants of E. coli MscL for their efficiency in disulfide bridging and generated a map of protein-protein interactions between these two regions. Interesting candidates have been further studied by patch clamp and show differences in channel activity under different redox potentials; the results suggest a model for the dynamics of these two domains during MscL gating.
  Channels (Austin, Tex.) 07/2012; 6(4). · 1.91 Impact Factor
• Article: The MscS and MscL families of mechanosensitive channels act as microbial emergency release valves.

  Ian R Booth, Paul Blount
  [show abstract] [hide abstract]
  ABSTRACT: Single-celled organisms must survive exposure to environmental extremes. Perhaps one of the most variable and potentially life-threatening changes that can occur is that of a rapid and acute decrease in external osmolarity. This easily translates into several atmospheres of additional pressure that can build up within the cell. Without a protective mechanism against such pressures, the cell will lyse. Hence, most microbes appear to possess members of one or both families of bacterial mechanosensitive channels, MscS and MscL, which can act as biological emergency release valves that allow cytoplasmic solutes to be jettisoned rapidly from the cell. While this is undoubtedly a function of these proteins, the discovery of the presence of MscS homologues in plant organelles and MscL in fungus and mycoplasma genomes may complicate this simplistic interpretation of the physiology underlying these proteins. Here we compare and contrast these two mechanosensitive channel families, discuss their potential physiological roles, and review some of the most relevant data that underlie the current models for their structure and function.
  Journal of bacteriology 06/2012; 194(18):4802-9. · 3.94 Impact Factor