An AFM study of solid-phase bilayers of unsaturated PC lipids and the lateral distribution of the transmembrane model peptide WALP23 in these bilayers

An altered lipid packing can have a large influence on the properties of the membrane and the lateral distribution of proteins and/or peptides that are associated with the bilayer. Here, it is shown by contact-mode atomic force microscopy that the surface topography of solid-phase bilayers of PC lipids with an unsaturated cis bond in their acyl chains shows surfaces with a large number of line-type packing defects, in contrast to the much smoother surfaces observed for saturated PC lipids. Di-n:1-PC (n = 20, 22, 24) and (16:0,18:1)-PC (POPC) were used. Next, the influence of an altered lipid environment on the lateral distribution of the single α-helical model peptide WALP23 was studied by incorporating the peptide in the bilayers of di-n:1-PC (n = 20, 22, 24) and (16:0,18:1)-PC unsaturated lipids. The presence of WALP23 leads to an increase in the number of packing defects but does not lead to the formation of the striated domains that were previously observed in bilayers of saturated PC lipids and WALP. This is ascribed to the less efficient lateral lipid packing of the unsaturated lipids, while the increase in packing defects is probably an indirect effect of the peptide. Finally, the fact that an altered lipid packing affects the distribution of WALP23 is also confirmed in an additional experiment where the solvent TFE (2,2,2-trifluorethanol) is added to bilayers of di-16:0-PC/WALP23. At 3.5 vol% TFE, the previous striated ordering of the peptide is abolished and replaced by loose lines

Developing a single-molecule fluorescence tool to quantify DNA damage

Quantification of DNA damage is an important technique for medical physics, for example to assess damage caused by the quinolone antibiotics, or to examine the effects of novel cancer treatments such as low-temperature plasma therapy on healthy or tumorous cells. Existing damage quantification techniques such as the alkaline comet assay [1] are often subjective in their results, especially at higher damage levels. Methods using immunofluorescence [2] often lose information due to the three dimensional nature of the cell

Imaging the proteins pseudoazurin and apo-pseudoazurin on gold by STM in air: effect of the bias voltage.

We have applied scanning tunnelling microscopy (STM) to the study of two proteins: pseudoazurin and apo-pseudoazurin. Both proteins adsorbed onto a Au (1 1 1) surface are visible to STM individually, forming into layers and multilayers, with currents from about 55 to 600 pA. The images reproduce well the expected dimensions laterally but not in the z direction. The apparent height of the proteins varies with the voltage polarity, being higher at negative sample voltages. The bias also affects their shape. Negative sample voltages of more than 1.5 V orient the proteins present on a gold terrace in parallel rows. The layer of water adsorbed on surfaces in ambient conditions can be related to our results to explain the reduced z dimensions, the asymmetry with the voltage polarity and the alignment of proteins at voltages more negative than -1.5 V

Ambient STM and in situ AFM study of nitrite reductase proteins adsorbed on gold and graphite: influence of the substrate on protein interactions.

Trimeric Achromobacter cycloclastes Cu-containing nitrite reductase (CuNIR) proteins adsorbed on gold and graphite have been studied by ambient STM and in situ AFM. STM resolves them individually and in layers, distinguishing the sub-molecular individual units of the trimer. The Cu atoms are not visible to STM. STM shows that individual CuNIR denatures as it adsorbs on Au, although a deformed trimeric shape can be identified in some cases. CuNIR forms disordered layers on gold. On graphite, ordered self-assembled layers of CuNIR have been resolved by in situ AFM and ambient STM forming parallel rows whose separation distance corresponds to the size of one of the units of the trimer, 5nm. Ambient STM can achieve better resolution than in situ AFM in the images of the layers. We observe differences between domains showing the parallel row structure and unstructured parts of the CuNIR layer by in situ phase imaging AFM

Scanning tunnelling microscopy images of the copper-containing amine oxidase from Arthrobacter globiformis in the holo and apo forms adsorbed on gold under ambient conditions

Dimeric Arthrobacter globiformis amine oxidase in the holo and apo forms adsorbed onto a Au (111) surface have been observed by scanning tunnelling microscopy (STM) under ambient conditions. Individual protein molecules denature as they adsorb onto a bare Au surface, although they keep a dual appearance. Tapping atomic force microscopy images of individual proteins correspond well with the STM ones in the lateral direction. STM voltage affects the distance between the units of the denatured proteins: negative voltages separate them while positive ones get them together. Disordered as well as ordered layers of apo and holo proteins have been resolved by STM at molecular detail level. Individual proteins lying on the layers present a compact and a distinct dimeric shape, apo dimers looking bigger than holo dimers

Lateral coupling and cooperative dynamics in the function of the native membrane protein bacteriorhodopsin

Membrane proteins are laterally coupled to the surrounding cell membrane through complex interactions that can modulate their function. Here, we directly observe and quantify the dynamics of functioning bacteriorhodopsin (bR) in its native membrane, a crystalline aggregate of bR trimers. We show that much of a monomer's isomerization energy is mechanically redistributed into the membrane, producing cooperative activity within the trimer while simultaneously generating functionally relevant long-range lateral pressure waves. Our results provide evidence of coordinated short and long-range effects in the cell membrane. © 2009 The Royal Society of Chemistry

Nanotribology of clean and oxide-covered silicon surfaces using atomic force microscopy

Atomic force microscopy (AFM) has been used for tribological studies of silicon surfaces both with and without an oxide layer on the surface. Three different types of surfaces were prepared: a silicon surface with a chemical oxide made by the SC1 process, a silicon surface with a thermal oxide, and a H-terminated silicon surface without an oxide layer. Only in the case of the chemical oxide, scratching of the oxide and ploughing of the silicon by the Si3N4 AFM tip were observed. On the other hand, no wear of the sample was noted on the other surfaces. On these surfaces, the AFM often produced elevated patterns in the shape of the scanned area, which were no longer visible after HF etching. The difference between the tribological behavior of the chemical-oxide-covered surface and that of the other surfaces is discussed in relation to the presence of hydroxyl groups in the oxide layer

Scanning tunneling microscopy study of the misfit layer compounds (LaSe)(x)NbSe2 and (PbSe)(x)NbSe2

Atomic images of the misfit layer compounds (LaSe)xNbSe2 and (PbSe)xNbSe2 were obtained with a scanning tunneling microscope (STM) operating in constant height mode in air. It was possible to record pictures of only the NbSe2 layers of both compounds, the LaSe and PbSe layers could not be observed. Formation of stage-2 portions embedded in the stage-1 crystal and instability of the LaSe and PbSe layers under the scanning conditions are discussed as possible causes. Comparing with the NbSe2 crystal, the lattice of the NbSe2 layers of the misfit layer compounds appear deformed. Nonperiodic Moiré-like structures have been observed in the (LaSe)xNbSe2 surface. We consider that this feature is caused by the STM tip pushing down the surface where it is softer. The nonperiodicity of the patterns might be due to the strain conditions of the crystal growth that would give rise to dislocations and defects