ORNL Biological and Nanoscale Systems research focuses on the characterization, integration and adaptation of natural and synthetic systems across multiple length scales. A continuing emphasis is to characterize and understand how natural systems are organized at the nanoscale and how this organization contributes to biological function.
To support this aim, an interdisciplinary group of researchers focuses on technology development with specific interests in biological imaging, biocompatible micro- and nanofabrication techniques, and high throughput screening technologies. Current imaging projects evaluate microbial systems and seek to trace the location and quantity of membrane proteins, to identify interacting proteins, and to confirm biochemical networks. Nanotechnology projects are centered on mimicking the physical and chemical characteristics of biological cells, developing biocompatible patterning techniques, and adapting biological routes to nanoscale material fabrication.
The group maintains comprehensive resources in molecular biology and in molecular and cellular imaging and leverages off of the advanced fabrication capabilities of ORNL’s Center for Nanophase Materials Sciences.
Understanding how organisms grow and respond to their environment requires the ability to interact with and manipulate biochemical pathways in live cells in real time. A comprehensive characterization of live cells involves knowing where proteins localize and how and when they interact with other proteins to form functional pathways. Imaging techniques are especially powerful because they can provide spatial and temporal localization data, as well as generate information about molecular interactions, chemical environment, and physical properties. Current research interests are focused on using genomics and live cell imaging tools to understand the function of proteins and networks in living cells.
Initial studies have focused on microbial pathways, including the glucose phosphotransferase system in E. coli. In order to visualize and track specific proteins in live cells, the proteins of interest must be labeled. One approach to protein labeling is through the use of recombinantly introduced peptide tags, such as green fluorescent protein (GFP) and its spectral derivatives. By genetically engineering protein fusions, perfect stoichiometry can be achieved, physiological responses can be measured in live cells, and additional labeling steps that might risk altering protein function are avoided.
Atomic force microscopy (AFM) is another imaging tool employed to provide a comprehensive analysis of protein networks. AFM allows molecular scale resolution of biological materials in physiologically relevant conditions and is capable of assessing the large proportion of the proteome that interacts at the cell membrane. In addition to imaging, the sensitivity of the AFM cantilever can be exploited to measure forces required to rupture bonds between biomolecules and within single protein molecules and to identify single molecule interactions with specific target molecules on surfaces by tethering specific probe molecules to the cantilever.
ORNL's nanotechnology research comprises several different projects, which are summarized below. These projects are directed towards designing, constructing and implementing nanoscale structures that are useful for either interfacing, mimicking or characterizing biological systems.
Nanosensing and Actuation Using Cell Mimetics
Functional biomedical sensing devices will need to directly interface to the molecular-scale processes of biology. These devices must extract and process information from a complex environment of interacting non-linear biomolecular processes and, in an ideal embodiment, intervene in processes gone awry. Functionality at this level of complexity dictates the requirements of engineering functional nanoscale components within microscale structures. Further, the research must employ processing schemes that are highly complex yet can be implemented in a very small volume. Nature’s answer to this design challenge is the cell, and the objective of this work is to develop sensors that mimic the dimensions and some portion of the functionality of a cell. Only by mimicking cellular features will effective operation and interfacing to biological systems be achieved. To achieve this ideal, ORNL researchers are exploiting recent advances in nanofabrication that allow for the synthesis of physical features on length scales ranging from nanometers to centimeters. These fabrication techniques allow for the construction of cellular mimetics that incorporate features such as semi-permeable membranes, chemical sensors and chemical actuators in a footprint of less than 100 µm. The nanostructured features are derived from the synthesis of carbon nanofibers that allow for control on the molecular scale. Researchers are establishing the functional elements of cellular mimics and integrating these capabilities for demonstrations of sensing and actuation at the molecular and cellular scale.
Molecular Scale Patterning of Biofunctional Surfaces via Scanning Probe Lithography
Small molecules, proteins, and cells are naturally subjected to interactions with surfaces that alter their physical and chemical makeup. These alterations are often dictated by the precise spatial arrangement of organic species within complementary molecular assemblies on the interacting surface. Investigation of the role that molecular scale assembly and arrangement of biofunctional surfaces have on fundamental cellular processes or subcellular biochemical reactions requires the capacity to engineer surfaces with different organic species at the nanoscale. To that end, researchers are integrating and developing multiscale, biocompatible lithography techniques based on microcontact printing, atomic force nanografting and related scanning probe lithography processes to engineer biofunctional surfaces with nanoscale precision. Current studies focus on the use of microcontact printing to develop micro- or cellular-scale patterns of biofunctional and inert alkanethiols on gold surfaces. These surfaces are characterized via closed loop, contact mode AFM scanning in a liquid environment and selectively modified at the nanoscale via physical removal, or “scraping”, using the AFM tip as a lithographic tool. These areas are simultaneously backfilled with biofunctional molecules present in the liquid imaging cell and tested for interactions with proteins and whole cells. These surfaces, having biofunctional elements that span multiple length scales, will not only facilitate a more detailed understanding of fundamental biological processes, but they will also find broad application in the fields of tissue engineering, drug delivery, biosensor design, and high throughput screening.
Nanoscale Devices for Biomolecular Interfaces
Small, engineered organic molecules are vital components of nanobiotechnological systems. One major role they play is as molecular-scale devices, where they represent the ultimate in miniaturization for nano-scale engineering. Researchers are particularly interested in light-operated devices that respond to light by undergoing molecular motion, such as azobenzene. Through appropriate chemical methods, azobenzene can be incorporated into more complex structures to couple this motion to downstream events, making the azobenzene a phototransducer. Researchers are focused primarily on incorporating azobenzene into amino acid- and peptide-based molecules for applications of using light to direct movement of proteins within cells, to stimulate neuronal signaling in the retina, and to control association of materials.
Additionally, small organic molecules have a valuable structural role in nanoscale devices, as they can both stabilize synthetic nanomaterials and provide the physical link to biomolecules, cells, or tissues. Researchers have recently initiated an effort to address the difficulty of forming stable nanoparticle-protein assemblies for biomedical applications.
For more Information
Biological & Nanoscale Systems Group
Oak Ridge National Laboratories
PO Box 2008 MS 6445
Oak Ridge, TN 37831-6445
Fax: (865) 574-5354