Concept of Size in Nanobiotechnology

Let us make an in-depth study of the concept of size in Nanobiotechnology. The below given article will help you to learn about the following things:- 1. The Overview on the Concept of Size in Nanobiotechnology 2. Molecules as Nanostructures and Molecules in Nanostructures 3. Molecular Machines and Organelles and 4. Micro/Nanostructures as Tools for Biotechnology.

The Overview on the Concept of Size in Nanobiotechnology:

The current enthusiasm for things ‘Nano’ in size has led naturally to a search for con­nections between these things and biology (the hottest area of science) and technology (where science pays off). Scientific interest in this intersection of fields is based on the perception that nanotechnology offers biol­ogy new tools, and that biology offers nano­technology access to new types of functional Nano systems—components of the cell—that are unquestionably interesting and possibly useful.

Public interest in the intersection of ‘Nano’ and ‘bio’ is also high. There are im­portant applications of Nano science in biol­ogy and biotechnology biology also provides unparalleled examples of functional nano­structures to excite the imagination of Nano-technologists of all persuasions.

The story is, however, not entirely ‘Nano’, but includes structures having a wide range of sizes. When small structures are considered for biologi­cal applications, or when small biologically derived structures are determined to have remarkable properties, the size of the system can be ‘Nano’ (which we define as 1-100 nm) but also ‘micro’ (from 100 nm, or 0.1 mm, to perhaps 1,000 µm, or 1 nm).

The ranges of sizes covered by these terms—Nano scale, micro scale and simply small are important. Structures vital to the cell have dimensions ranging from those of small molecules to these of millimeter-scale fluidic devices which size is most important depends on what ques­tion one is asking. Enthusiasm for the poten­tial value of ‘Nano’ should be balanced against the established and rapidly expanding value of ‘micro’.

Nano science and nanotechnology have grown exuberantly in a rich mixture of le­gitimate scientific opportunity, technological imperative and hyperbole. Much of the initial impetus for the development of nanotechnol­ogy came from its relevance to electronics. Microelectronics has, unarguably, changed the world through its impact on informa­tion processing and communications, and the progress in the field has been measured by its adherence to Moore’s Law. As micro­electronic devices have become smaller, they have become less expensive, faster and more portable.

Until recently, the critical lateral di­mensions in integrated circuits were greater than 100 nm will also be an important tech­nology. The majority of the very small devices used in Nano electronics will be generated at least initially by evolutionary technologies devel­oped from the existing methods of micro­electronics (that is, methods of fabrication based primarily on photolithography).

It is not clear yet whether revolutionary tech­nologies based, perhaps, on the remarkable electron transport properties of silicon Nano rods, or on nanometer structures supporting quantum computation, or on scanning problem microscopies for ultra-dense information storage will emerge as sufficiently practical to be commercialized.

New catalytic processes for growth of materials make nanotubes and Nano spheres readily available in a wide vari­ety of compositions. Electron beam writing has begun to move into the mainstream of fabrication of nanostructures, and electron microscopy is, of course, a mainstay of ob­servational Nano science.

The situation in bio­logical science is different for several reasons:

First, biological structures are relatively large compared to structures in electronics and in physical Nano science. A mammalian cell is approximately 10 µm in a diameter when rounded, and, perhaps, 50 µm in diameter when fully spread in attached tissue culture— these dimensions will not change.

Although it is certainly important to explore the small­er, biologically vital components of the cell, the technological imperative to make things smaller that has dominated microelectronics does not have the same urgency in biology. The ability to observe intracellular structures with high selectivity, and to follow the dynamic behaviour of these structures, may be more important than the ability simply to resolve small features by some form of microscopy.

Second, the scanning probe microscope has been less revolutionary in its impact on biol­ogy than on the physical sciences. Biological structures even those on surfaces are soft and electrically insulting, and not easily imaged. Moreover, most of biology goes on inside the cell, where the scanning probe tip cannot reach.

Third, there already exists a highly de­veloped science concerned with biologically relevant nanostructures: ‘biochemistry’ and it fabricates biological nanostructures of all sizes, from low-molecular weight drugs to DNA—with dimensions from a nanometer to hundreds of microns.

The Cell is the Core of Biology:

It is the smallest unit that is alive. The cell is, in a reductionist view, a compartment in which a collection of reactions occurs. These reac­tions, essentially all catalysed, modulate one another, and together from a network with the remarkable properties like self-replication, energy dissipation, adaptation etc. and form what we call ‘life’.

The scale of sizes that characterize chemical bonds, and the mole­cules and even very large molecules made out of simple molecules, is familiar. The cell is, in a holistic view, an entity with phenotypic properties and behaviours; it moves, repli­cates or destroys itself, harvests energy from sunlight or generates it in useful form by burning glucose and di-oxygen, applies force, transmits signals, senses its environment, stores and transmits information.

The science of biological molecules studied ex vivo, in di­lute buffer, is highly advanced; the science that studies higher-order behaviours of cells (and of molecules in cells) is still developing. There would, arguably, be no need to invent a new discipline of Nano biology, were there not concepts and tools that were not adequately covered by the existing disciplines. In fact, there are many unmet needs.

Molecules as Nanostructures and Molecules in Nanostructures:

Conventional molecular science (in the guise of enzymology, or analytical biochemistry, or biophysics) deals competently with molecules in solution. In the cell, however, molecules are often organized into functional aggre­gates, normally with nanometer scale dimen­sions.

Visualizing and studying these struc­tures especially as they change dynamically during cycles of function is one of the key challenges posed to Nano science by biology. Progress toward these objectives is now rap­id: spectroscopic studies of single molecules provide one example; biophysical studies of multi-protein aggregates are a second.

Single Molecules:

The single molecule is, in a sense, the ultimate nanostructure. The average behaviour of a single molecule, ob­served over a long period of time, is the same as the behaviour of a collection of molecules. The sensitivity of analytical methods has been such that only their ensembles could be stud­ied.

The rapidly growing ability (based on advances in both lasers and detectors) to ob­serve single molecules using high-resolution microscopy, and to examine the fluctuations in the behaviour of these molecules over short times, is revealing a wealth of new in­formation about the dynamics of molecules, especially catalyst, relevant to biology.

A substantial effort is focused on observa­tions of catalytic activity in enzymes, where at least some of the fluctuations are relatively slow. It is not presently clear whether these fluctuations are biologically functional, but the possibility that they might contribute to the control of metabolic or signal-transduction pathways is a new element to consider in systems biology.

Single-molecule microscopy is one of a number of new optical techniques that are either circumventing the classical limits of microscopy or providing dramati­cally more informative images within the classical limits. Confocal microscopes, near field optical systems, highly sensitive Raman microscopes and total internal reflection mi­croscopes provide further examples of the rapid progress being made in characterizing nanostructures using light microscopy.

Molecular Machines and Organelles:

The ultimate in functional Nano systems ‘biological Nano machines’ populate the cell. The ribosome, Na⁺/K⁺ ATPase, flagellar mi­cro-motor of bacteria, linear micro-motors of muscle and of the microtubules that organize and move the cell, voltage-gated ion chan­nel. DNA replication complexes, multimeric membrane receptors, and the photosynthetic reaction center; these, and countless other structures in the cell, are astonishingly com­plex, non-intuitive in their operation, and instructive to contemplate.

Path clamping was one of the first techniques for monitoring the activity of single molecules and small protein complexes. Information generated by X-ray crystallography has begun to clarify these nanostructures. We can see that their func­tion depends centrally on the catalytic activi­ties of their constituent proteins, on the modulation of these activities by changes in the conformation of the proteins induced by the environment (e.g., the trans-membrane po­tential) and by changes in the conformation reflecting catalysis.

No technique yet has the sensitivity and resolution to allow the direct observation of single molecules during com­plete cycles of biological activity, but combinations of crystallography carried out on intermediates in these cycles with other kinds of information are beginning to clarify what happens during functional catalytic cycles or signaling events; understanding of why these cycles proceed as they do (in terms of the details of protein structure, organization and energetics) lies ahead.

Understanding biological nanostructures will be enormously stimulating for nanotechnology. The concept of very tiny machines has always appealed to scientists and non-scientists alike but the na­scent field of nanotechnology originally as­sumed that Nano machines would probably resemble macro machines in their design.

It is seldom possible to prove that something cannot happen in science and it is difficult to prove that one cannot build Nano machines that are analogous to familiar macro machines. The fact that biological structures, although functioning in familiar ways, operate using principles that are entirely unfamiliar based on everyday experience suggests that would- be designers of Nano machines have much to learn from biology.

An example is the rotary flagellar motor of bacteria (and the function­ally related Na⁺K⁺ ATPase). This motor does have a structure that serves as a shaft, and another structure that anchors the motor in the cell membrane, but, beyond that, any re­semblance to a macroscopic motor (whether internal combustion or electrical) stops.

The components of the machine are complex, three-dimensional structures (proteins) that self-assemble in a series of steps, starting from a linear polypeptide chain; the mecha­nism of rotary motion seems to involve elec­trical current, magnetic fields or expansion of hot gases in a cylinder.

Micro/Nanostructures as Tools for Biotechnology:

One of the areas where biology can ben­efit from Nano science is new materials and structures. There is a wide variety: surfaces patterned with self-assembled monolayers (SAMs) to guide cell attachment and growth; needles, holes and channels for biophysical tools; microstructure, three-dimensional scaffolds for tissue engineering; nanoparticles as probes; photonic band-gap structures; new optical systems for imaging.

There are many such examples which represent a wave of de­velopment of tools by physical science for ap­plication to biological science, and are a part of the rapprochement between the physical and biological sciences. A few examples illus­trate these rapidly developing areas.

Self-Assembled Monolayers (SAMs):

SAMs have provided the ability to ‘synthesize’ extended surfaces. Dipping a thin gold film (typically 40 nm thick, supported on glass or silicon) into a solution of an alkanethiol [R(CH₂)₁₁–₁₈SH] forms SAMs. The sulfur atom chemisorbs on the gold releases a hy­drogen atom and forms a strong sulfur-gold bond.

This process allows the ‘synthesis’ of macroscopic structures comprising areas of square centimeters of ordered molecules. These structures typically contain ~5 x 10¹⁴ molecules/cm² and are a form of polymer; a structure containing many organic side groups (the thiols) attached covalently to a backbone (the gold film).

They comprise numbers of atoms that could not be ‘synthe­sized’ in any conventional approach to organ­ic synthesis. These surfaces are needed for ap­plications in cell biology, as the projected area of a cell onto a surface of this type includes ~ 10⁹ – 10¹⁰ thiolate molecules.

SAMs are nano­structures materials-interfacial films with a thickness of approximately 2 nm, which al­low atomic-level precision (relative to the mean plane of the surface) in the placement of functional groups. As the structures self-assemble, relatively simple synthesis of the precursors allows the formation of structures presenting complex, functional ligands.

An example demonstrates the versatility of SAMs in controlling the environment of cells. SAMs terminating in methyl groups are hydrophobic, and adsorb proteins from solution. SAMs terminating in oligo (ethyl­ene glycol) moieties resist the adsorption of proteins from solution.

When cells attach to a surface, they do not, in general, attach to the surface directly, in fact they attach to proteins adsorbed on the surface. The combination of SAMs with ‘printing’ using elastomeric stamps (so-called ‘soft lithography’) allows the surface to be patterned into regions to which cells attach and regions to which they do not.

Having allowed the cells to attach and to spread to the limits of the regions to which they adhere, it is then possible to employ a useful electrochemical trick developed in sur­face science for other reasons, for example, application of a brief voltage pulse to the gold layer selectively detaches the non-adsorbing thiols.

As soon as they have left the surface, proteins from the culture medium adsorb on the gold, and generate a surface across which cells can spread. The ability to grow cells in patterns, and then to release them from these patterns with a simple electrochemical ma­nipulation not to damage them, provides the basis for new types of bioassays that make use of observations of cell motility.

Nano Tips and Microspheres:

The origin of nanotechnology is tools for im­aging nanostructures; originally electron mi­croscopy, but, more recently, and famously, scanning probe devices. Although the direct imaging of biological structures has proved difficult, scanning probe devices have been enormously successful in allowing the appli­cation of forces directly to single molecules.

The kind of information provided by these force-distance curves has, for example, made it possible to infer how stress is stored as strain in molecules by the unfolding of pro­tein domains. Complementary information comes from studies in which the force is ap­plied using optical tweezers or, more recently, magnetic beads.

The use of optical tweezers has provided insights into the mechanism and function of proteins involved in active transporting the cells (e.g., myosin and kinesin). Magnetic tweezers have the singular advantage that it is possible to carry out paral­lel measurements on large numbers of beads, and thus to improve the statistics of measure­ments.

Channels and Pores:

Micro channels are the basis of microfluidic devices. When micro channels were made in silicon using conventional photolithography, the techniques were too slow, specialized and cumbersome to be broadly useful to biologists. The development of soft lithographic meth­ods for micro fabrication, and the realization that high resolution printing provided ad­equate resolution for fabrication of channels with widths from 10 µm to 1 mm, opened the door to active development of this area as a convenient and practical technique for fabricating microfluidic devices.

The physics of fluids flowing in micro channels provided another important component of this area of micro technology. In micro channels, aqueous buffers almost always flow luminary—that is, without eddies, and with only diffusive mix­ing of streams flowing side by side. This com­bination of physical phenomena has begun to generate a wide variety of new devices: micro cell separators and particle counters, micro­systems for cell culture, gradient generators and analytical systems.

Using more complex methods of fabrication, it is now possible to make true Nano channels. The practical value of these systems remains to be seen as it is difficult to keep them from plugging up with particles invariably present in real systems, but they do have the advantage that their di­mensions are smaller than the size of biologi­cal macromolecules, with potentially useful consequences for separations; they also have a very high ratio of surface to volume, and thus allow the study of wall effects in biologi­cal separations.

Micro and nanofabrication techniques also offer access to pores (which are, in essence, very short channels). Nano-fabricated pores with dimensions down in approximately 2 nm have been demonstrated and proposed as the basis for single-molecule DNA sequencing.

Although it is uncertain whether this application will ultimately suc­ceed, pores will certainly be useful in a range of other applications. Interestingly, biology is beginning to offer its own set of methods for making channels and pores with Nano scale dimensions; the case with which some of these systems can be assembled suggests that they may, in the long run, provide systems that are at least as useful as those fabricated top-down. Lipid nanotubes and channels in lipid membranes based on pore-forming peptides and proteins are examples.

Nanoparticles:

Nanoparticles are among the first Nano scale materials to be directly useful in biology. Fluorescent particles labeled with antibod­ies (as tags that do not photo bleach) and super-paramagnetic magnetite particles coated with dextran (as image enhancement agents in magnetic resonance imaging) are commer­cially available; a wide range of other fluores­cent or magnetic nanoparticles will soon be available.

Eventually, these small particles must be made much more useful and infor­mative if they are to play an important role in understanding the workings of the cell, but nanotechnology has clearly identified the field of nanoparticles as one where new tech­niques in synthesis will make a wide range of particles available, and where these particles meet a need as labels for biological structure and function.

To the physical science, ‘Nano’ offers quantum phe­nomena (size-dependent fluorescence, long ballistic electron trajectories) and remark­able physical properties (mechanical moduli, electrical conductivity).

Biology adds incred­ibly sophisticated Nano machines, operating by entirely classical molecular mechanism. To the biological sciences, ‘Nano’ offers new tools, many from the physical sciences, that will be necessary to put together a conceptual model of life, and a fresh framework on which to hang ideas about functional aggregates (i.e., biological Nano machines).

The road from reading the information in the genome to un­derstanding life will be an arduous one, and reading the genome may be the easiest part. Understanding how molecules organize and function in cells will require new tools and concepts, and, as the assembles of molecules of greatest interest will have nanometer di­mensions, the tools must be appropriate for the task, that is, they must be able to char­acterize structures 0.1-0.001 times the size of the cell. At the same time, selecting, sorting, maintaining, stimulating, herding and char­acterizing the cells will require tools substan­tially larger than the cell. Both nanometers and micrometer dimensions are relevant.

The different perspectives are not quite as dispa­rate as those of the seven blind men with an elephant (or perhaps a flea, in this context), but the idea remains. Nano and micro science will show different aspects to different fields, and the integration of these aspects will yield some of the new concepts and techniques that will build a more complete picture of the cell.

The flagellar rotary motor again provides an example of the range of opportunities and dimensions facing ‘Nano biotechnology’. This structure might provide an illustration of ‘principles’ that could be used to design a non-protein nanometer; it might be useful left in vivo, with the organism itself employed to do the work. Each application has possibili­ties and each involves different scales of siz­es, and different critical dimensions. ‘Small’ —both ‘Nano’ and ‘micro’—must be a part of the future of biotechnology.