David CafisoDavid S. Cafiso

  • Professor of Chemistry

  • A.B., University of California, Berkeley, 1974
  • Ph.D., University of California, Berkeley, 1979
  • Postdoctoral Fellow, Stanford University, 1980

  • Jane Coffin Childs Foundation Fellowship, 1979-1981
  • Camille and Henry Dreyfus Foundation Award
    for New Faculty in Chemistry, 1981

  • Phone: (434) 924-3067
  • Cafiso Lab Group Website

Biophysical Chemistry; Membrane Proteins and Cell Signaling

Membranes and membrane proteins participate in some of the most important and interesting cellular processes. Energy transduction, cell signaling, membrane excitability, secretion and immune recognition are examples of a few of the processes mediated by membrane proteins. However, the molecular mechanisms by which lipids and membrane proteins accomplish these tasks are largely unknown. Approximately 30 to 40 percent of the genome codes for membrane proteins and 70 percent or more of current pharmaceuticals appear to act on membrane proteins. For these reasons, membrane proteins represent one of the most important challenges in the field of structural biology.

Work in our laboratory is directed at studying membranes and membrane proteins, and there are currently two general areas of ongoing research. One area of investigation involves studies on the mechanisms by which proteins become attached to membrane surfaces. Attachment is critical for cell-signaling because it controls protein-protein interactions and the access of enzymes to lipid substrates. For example, the oncogenic form of the src tyrosine kinase is not active and fails to transform cells until it becomes attached to the cytoplasmic face of the plasma membrane. We are currently determining the structure and electrostatic interactions made by highly positively charged protein motifs, such as those from MARCKS (the myristoylated alanine rich C-kinase substrate) with negatively charged lipid surfaces. In addition to regulating membrane attachment, these positively charged motifs function to sequester phosphatidylinositol 4,5, bisphosphate (PIP2), and regulate the activity of this phosphorylated inositol lipid within the cytoplasmic membrane. We are also determining the membrane interactions made by protein domains such as C2 domains, which are found in a wide range of proteins involved in cell-signaling. C2 domains function to attach their parent proteins to membranes in a Ca ++ dependent fashion. C2 domains perform critical roles in membrane trafficking, membrane fusion and membrane repair, and defects in these domains result in forms of muscular dystrophy and deafness.

A second area of investigation involves membrane transport. We are currently examining the molecular mechanisms that function to facilitate active transport. For example, we are determining the molecular mechanisms by which BtuB transports vitamin B12 across the outer membrane of Escherichia coli. This protein is homologous to FecA, FepA and FhuA, outer membrane iron transport proteins that presumably function by similar mechanisms. These proteins belong to a class of transport proteins for which high-resolution structural models have been obtained, and they are extremely important for the survival of some bacterial pathogens. In addition to BtuB, FecA and FhuA, we are presently expressing, reconstituting and labeling BtuC/D. This protein is member of the ABC cassette transporter family and it is responsible for carrying vitamin B12 across the inner membrane.

The primary tools that we use in our work include EPR spectroscopy and high-resolution NMR. Site-directed spin labeling is a powerful methodology that combines site-directed mutagenesis with and EPR spectroscopy. We are developing and making use of this tool, which is particularly well-suited to address questions regarding the dynamics and molecular function of membrane proteins.

Representative Publications

Membrane Bound Orientation and Position of the Synaptotagmin C2B Domain Determined by Site-Directed Spin Labeling. E. Rufener, A. Frazier, C. M. Wieser, A. Hinderliter, and D. S. Cafiso. Biochemistry, 44, 18-28 (2005).

Membrane Position of a Basic Aromatic Peptide that Sequesters Phosphatidylinositol 4,5 Bisphosphate Determined by Site-Directed Spin Labeling and High-Resolution NMR. J. F. Ellena, J. Moulthrop, J. Wu, M. Rauch, S. Jaysinghne, J. D. Castle, and D. S. Cafiso. Biophysical Journal, 87, 3221-3233 (2004).

Spectroscopic Evidence that Osmolytes Used in Crystallization Buffers Inhibit a Conformation Change in a Membrane Protein. G. E. Fanucci, J. Y. Lee and D. S. Cafiso. Biochemistry 42, 13106-13112 (2003).

Membrane Mimetic Environments Alter the Conformation of the Outer Membrane Protein BtuB. G. E. Fanucci, J. Y.Lee and D. S. Cafiso. in the press JACS (2003).

Substrate-Induced Conformational Changes of the Perplasmic N-Terminus of an Outer-Membrane Transporter by Site-Directed Spin Labeling. G. E. Fanucci, K. A. Coggshall, N. Cadieux, M. Kim, R. J. Kadner and D. S. Cafiso. Biochemistry 42, 1391-1400 (2003).

Membrane bound orientation and position of the synaptotagmin I C2A domain by site-directed spin labeling. A. A. Frazier, C. R. Roller, J. J. Havelka, A. Hinderliter and D. S. Cafiso. Biochemistry, 42, 96-105 (2003).

Structure and Dynamics of the b-Barrel of the Membrane Transporter BtuB by Site-Directed Spin Labeling. G. E. Fanucci, N. Cadieux, C. A. Piedmont, R. J. Kadner, D. S. Cafiso. Biochemistry ; 41, 11543-11551 (2002).

Peptide-membrane interactions determined using site directed spin labeling. D. S. Cafiso. Current Topics in Membranes 52, 3-29 (2002).

Membrane orientation and position of the C2 domain from cPLA2 by site-directed spin labeling. Frazier, A. F., Wisner, M. A., Malmberg, N. J., Victor, K. G., Fanucci, G. E., Nalefski, E. A., Falke, J. J., Cafiso D. S. Biochemistry 41, 6282-6292 (2002).

Myristoylated Alanine-rich C Kinase Substrate (MARCKS) Sequesters Spin-labeled Phosphatidylinositol 4,5-Bisphosphate in Lipid Bilayers. Rauch, M. E., Ferguson, C. G., Prestwich, G. D. and Cafiso, D. S. J. Biol. Chem. 2002. 277: 14068-14076 (2002).

Location and Dynamics of Basic Peptides at the Membrane Interface: Electron Paramagnetic Resonance Spectroscopy of Tetramethyl-Piperidine-N-Oxyl-4-Amino-4-Carboxylic Acid-Labeled Peptides. K.G. Victor and D. S. Cafiso. Biophys. J. 81, 2241-2250 (2001).

Transport-Defective Mutations Alter the Conformation of the Energy-Coupling Motif of an Outer Membrane Transporter. K. A. Coggshall, N. Cadieux, C. Piedmont, R. J. Kadner, D. S. Cafiso. Biochemistry 40, 13964-13971 (2001).

Monitoring Conformational Changes with Site Directed Spin Labeling.  W. L. Hubbell, D. S. Cafiso, and C. Altenbach.  Nature Struct. Biol. 7, 735-739 (2000).

Substrate-induced exposure of an energy-coupling motif of a membrane transporter.  H. Merianos, N. Cadieux, C. Lin, R. Kadner, D. Cafiso.  Nature Struct. Biol.  7, 205-209 (2000).

Interactions Controlling the Membrane Binding of Basic Protein Domains.  Phenylalanine and the Attachment of the MARCKS Protein to Interfaces.  K. Victor, J. Jacob and D. S. Cafiso.  Biochemistry 38, 12527-12536 (1999).

Membrane Spontaneous Curvature Modulates the Binding Energy of a Channel Forming Voltage-Gated Peptide.  Lewis, J. R. and D. S. Cafiso.  Biochemistry 38, 5932-5938 (1999).

The Role of Proline and Glycine in Determining the Backbone Flexibility of a Channel Forming Peptide.  Jacob, J., Duclohier, H. and D. S. Cafiso. Biophysical Journal  76, 1367-1376 (1999).

Structure and Position of the N-terminal Binding Domain of pp60src at the Membrane Interface.  Victor, K., and D. S. Cafiso.  Biochemistry 37, 3402-3410 (1998).

Dipole potentials and spontaneous curvature:  membrane properties that could mediate anesthesia.  David S. Cafiso.  Toxicology Letters, 103, (1998).

Structural Features that Modulate the Transmembrane Migration of a Hydrophobic Peptide in Lipid Vesicles.  Jayasinghe, S., Barranger-Mathys, M., Ellena, J. F., Franklin, C., and D. S. Cafiso.  Biophysical J . 74, 3023-3030 (1998).

MARCKS, Membranes, and Calmodulin: Kinetics of Interaction.  Arbuzova, A., Wang, J. Murray, D., Jacob, J., Cafiso, D., McLaughlin, S.  Journal of Biological Chemistry.  272, 27167-27177 (1997).

Estimating the Electrostatic Potential at the Acetylcholine Receptor Agonist Site Using Power Saturation EPR.  George H. Addona and David S. Cafiso. BBA Biomembranes. 1329, 74-84 (1997).

Contrasting Membrane Localization and Behavior of Halogenated Cyclobutanes that Follow or Violate the Meyer-Overton Hypothesis of General Anesthetic Potency.  C. L. North and D. S. Cafiso  Biophys. J.  72, 1754-1761 (1997).

Defining protein-protein interactions using site-directed spin-labeling:  the binding of protein kinase C substates to calmodulin.  Qin, Z, Wertz, S. L., Jacob, J., Savino, Y. and D. S. Cafiso  Biochemistry 35, 13272-13276 (1996).

Solution and membrane bound structure of a peptide derived from the protein kinase C substrate domain of neuromodulin.  Wertz, S. L. Savino, Y. and D. S. Cafiso  Biochemistry 35, 11104-11112 (1996).

Membrane  Structure of the PKC and Calmodulin Binding Domain of MARCKS Determined by Site-Directed Spin-Labeling.  Q, Zhihai, and D. S. Cafiso Biochemistry, 35, 2917-2925 (1996).

Membrane Structure of Voltage-Gated Channel Forming Peptides by Site-Directed Spin-Labeling.  M. Barranger-Mathys and Cafiso, D. S.  Biochemistry, 35, 498-505 (1996).

Membrane Orientation of the N-terminal Segment of Alamethicin Determined by Solid-State 15N NMR.  C. L. North, M. Barranger-Mathys and D. Cafiso, Biophysical Journal 69, 2392-2397 (1995).

Distribution of General Anesthetics in Phospholipid Bilayers Determined Using 2H NMR and 1H-1H NOE Spectroscopy.  James Baber, Jeffrey F. Ellena, David S. Cafiso.  Biochemistry 34, 6533-6539 (1995).

Anesthetics Reduce the Magnitude of the Membrane Dipole Potential.  Measurements in Lipid Vesicles Using Voltage-Sensitive Spin-Probes.  Zhihai Qin, Gabor Szabo, and David Cafiso Biochemistry 34, 5536-5543 (1995).

The Structure of Micelle Associated Alamethicin from 1H NMR.  Evidence for Conformational Heterogeneity in a Voltage-Gated Peptide.  J. C. Franklin, J. F. Ellena, S. Jayasinghe, L. P. Kelsh and D. S. Cafiso.  Biochemistry 33, 4036-4045 (1994).

Collisions Between Helical Peptides in Membranes Monitored Using Electron Paramagnetic Resonance.  Evidence that Alamethicin is Monomeric in the Absence of a Membrane Potential.  M. Barranger and D. S. Cafiso.  Biophysical Journal 67, 172-176 (1994).

Molecular Flexibility Demonstrated by Paramagnetic Enhancements of Nuclear Relaxation.  Application to Alamethicin, a Voltage-Gated Peptide Channel.  C. L. North, J. C. Franklin, R. G. Bryant, and D. S. Cafiso.  Biophysical Journal 67, 1861-1866 (1994).

Alamethicin: a Peptide Model for Voltage-Gating and Protein-Membrane Interactions.  D. S. Cafiso  Annual Review of Biophysics and Biomolecular Structure  23, 141-165 (1994).