How do you determine shape and size of biomolecule in solution?

Answer 1

Size is easy. There are a number of techniques, some more accurate than others:

Mass spectroscopy - very accurate (to within ~0.1%). Requires a solution of about 1mg/ml.

Gel filtration chromatography - not particularly accurate (to within ~10%) and also influenced by shape.

SDS-PAGE - very crude estimation for proteins (commonly greater than 10% error) but based solely on molecular weight as molecules are denatured and given uniform charge.

Agarose gel - rough estimation for DNA.

Sequencing - protein/DNA sequencing would allow calculation of an approximate size, but it is much easier to use mass spectroscopy.

Analytical centrifugation - accurate but takes a very long time so rarely used.

A combination of the above methods can provide information on shape too. A typical early experiment on a protein, for example, might include both SDS-PAGE and gel filtration. Since one depends on size and shape, while the other is only size, one can determine its oligomer state (whether or not it binds to itself in solution) if it is known to be globular, or it can indicate if a protein is not roughly spherical (most are).

Shape is more complex:

As discussed above some basic information on shape (linear or globular and oligomerization) can be gained from the simple methods listed above. Further information can be gained through X-Ray crystallography, cryoelectron microscopy or COESY/NOESY 3D NMR spectroscopy. One of these techniques is usually able to give a reasonable high resolution three dimensional structure of a protein, but it is a time consuming and costly process.

For the very small biomolecules, mass spectroscopy and NMR spectroscopy (but not NOESY/COESY) are usually sufficient for determining both size and shape, as shape can be predicted using known bond length and angle constraints to a reasonable degree of accuracy.