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Faraday Discuss., 1992, 94, 183-197

Scanning Tunnelling Microscopy Observations of Biomolecules on Layered Materials Helmut Jungblut," Sheelagh A. Campbell,b Michael Giersig," Daniel J. Muller" and Hans Joachim Lewerenz" a

Hahn-Meitner-Institut Berlin, Bereich Photochemische Energieumwandlung, Postfach 39 01 28, D-1000 Berlin 39, Germany Portsmouth Polytechnic, Department of Chemistry, St. Michaels Building, White Swan Road, Portsmouth, UK PO1 2DT

Scanning tunnelling microscopy (STM) has been performed on the reverse transcriptases of the human immunodeficiency virus (HIV-1) and the moloney murine leukaemia virus (MuLV). The biological molecules are adsorbed on n-type semiconducting MoTe,. The p66 (66 kD) subunit of the RT of HIV-1 is imaged by STM. Both STM and processed transmission electron microscopy (TEM) data show a spherical and horseshoe-like shape of external diameter ca. 65 A, depending on the angle of observation. The STM results show a larger diameter which is related to the curvature radius of the tip of the probing needle. The RTs of HIV-1 and MuLV exhibit a circular hole of ca. 20 8, diameter in accordance with structure predictions and functioning considerations. The surface-molecule interaction is discussed in terms of the electronic properties of the semiconductor surface including the influence of small defect sites at the layered crystal surface.

1. Introduction The potential applicability of STM for the imaging of macromolecules and, in particular, biological molecules, was recognized almost immediately after the introduction of the technique.'.* The initial investigations focused on the possibilities of reproducing the structure of DNA m01ecules.~*~ Most of the results obtained needed substantial data processing in order to provide STM images which could be compared with theoretical model^.^'^ Layered graphite substrates have been almost exclusively used in these studies and various procedures which might have seriously affected the biomolecules have been to improve the bonding interaction at the substrate surface. In this work we depart from graphite as a substrate for imaging of biological molecules and explore the potential of Group VI layered transition-metal dichalcogenides for this function. Among this class of compounds, the semiconductor MoTe, has been chosen for the following reasons: first, the top of the valence band is of less pure metallic d-band character than that of the sulfide and selenide analogues.' Therefore the increased hybridization of the chalcogenide 5p orbitals with metal d-orbitals might result in a more directed bonding interaction at the outmost surface layer than with MoS, or MoSe, , for instance, which exhibit more ideal van der Waals behaviour." Secondly, it is possible to produce defects of varying size on the surface of MoTe, by cleaving and by the needle-substrate interaction in an STM experiment.' Such defects, at which strong covalent metal d-type bonds are exposed to the ambient, might act as specific bonding centres for large molecules and might leave their lateral shape largely unaffected, provided the defect site is much smaller than the adsorbed molecules. 183

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STM on Biomolecules

The necessity to understand, and possibly interrupt, the life cycle of retroviruses has led us to investigate a key enzyme in the viral reproduction process, the reverse transcriptases (RTs).'~ The main feature of RTs is their expression of viral RNA into double-stranded DNA which can then be integrated into the host-cell genome." Since the recognition of the function of RT more than two decades ago,14,15a wealth of information on these molecules has been accumulated, including structure predictions, l 6 > l 7DNA-synthesis m e c h a n i ~ m ' and ~ ~ 'recently ~ the first experimental information.*' So far, however, a direct imaging of these molecules has only been communicated in very recent reports from our In the present article, we present new experimental data on the RT of the human immunodeficiency virus (HIV-1) and of the moloney murine leukaemia virus (MuLV). The STM results on the former RT are substantiated by combination with high-resolution transmission electron microscopy (TEM),23structure predictions16 and more indirect experimental results from small-angle neutron scattering.*'

2. Experimental Instrumental Methods STM experiments were carried out using a Nanoscope I1 (Digital Instruments Inc.), fitted with Pt-Ir tips, supplied by Material Analytical Services. The needles were guaranteed to have tip radii smaller than 50nm, but examination of several of these needles, using scanning electron microscopy, showed great variations in tip radii up to 400nm. As improved images were obtained with needles possessing smaller tip radii, tip selection, as will be explained in section 3.3, was an important factor in obtaining high-resolution micrographs. Therefore, only needles with radii

Scanning tunnelling microscopy observations of biomolecules on layered materials.

Scanning tunnelling microscopy (STM) has been performed on the reverse transcriptases of the human immunodeficiency virus (HIV-1) and the moloney muri...
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