Molecular-level studies of extracellular matrix proteins conducted using atomic force microscopy.

Extracellular matrix (ECM) proteins provide anchorage and structural strength to cells and tissues in the body and, thus, are fundamental molecular components for processes of cell proliferation, growth, and function. Atomic force microscopy (AFM) has increasingly become a valuable approach for studying biological molecules such as ECM proteins at the level of individual molecules. Operational modes of AFM can be used to acquire the measurements of the physical, electronic, and mechanical properties of samples, as well as for viewing the intricate details of the surface chemistry of samples. Investigations of the morphology and properties of biomolecules at the nanoscale can be useful for understanding the interactions between ECM proteins and biological molecules such as cells, DNA, and other proteins. Methods for preparing protein samples for AFM studies require only basic steps, such as the immersion of a substrate in a dilute solution or protein, or the deposition of liquid droplets of protein suspensions on a flat, clean surface. Protocols of nanolithography have been used to define the arrangement of proteins for AFM studies. Using AFM, mechanical and force measurements with tips that are coated with ECM proteins can be captured in ambient or aqueous environments. In this review, representative examples of AFM studies are described for molecular-level investigations of the structure, surface assembly, protein-cell interactions, and mechanical properties of ECM proteins (collagen, elastin, fibronectin, and laminin). Methods used for sample preparation as well as characterization with modes of AFM will be discussed.

Structural Information on Supramolecular Copper(II) ß-Diketonate Complexes from Atomic Force Microscopy and Analytical Ultracentrifugation.

Supramolecular Cu(II) complexes were prepared from two trifunctional ß-diketone ligands. The ligands (CH3Si(phacH)3 and CH3Si(phprH)3, represented by LH3) contain three aryl-ß-diketone moieties joined by an organosilicon group. The complexes have the empirical formula Cu3L2, as expected for combinations of Cu2+ and L3-. Several metal-organic polyhedra (MOPs) [Cu3L2]n are possible (n = 1-10); a dodecahedron (Cu30L20; n = 10; estimated diameter of ca. 5 nm) should be the most stable because its internal bond angles would come closest to ideal values. Atomic force microscopy (AFM), performed on samples deposited from solution onto mica substrates, revealed a distribution of sample heights in the 0.5-3.0 nm range. The most commonly observed heights were 0.5-1.5 nm, corresponding to the smallest possible molecules (Cu3L2, i.e., n = 1). Some molecular cubes (Cu12L8; ca. 2.5 nm) or larger molecules or aggregates may be present as well. Equilibrium analytical ultracentrifugation (AUC) was also used to probe the compounds. A previously reported reference compound, the molecular square Cu4(m-pbhx)4 (M = 2241 g mol-1), behaved well in AUC experiments in four nonpolar organic solvents. AUC data for the new tris(ß-diketonate) MOPs [Cu3L2]n in toluene and fluorobenzene did not agree well with the theoretical results for a single solute. The data were fit well by a two-solute model, but these results were not consistent in the two solvents used, and some run-to-run variability was noted even in the same solvent. Also, the calculated molecular weights differed significantly from those expected for [Cu3L2]n ([Cu3(CH3Si(phac)3)2]n, multiples of 1322 g mol-1; or [Cu3(CH3Si(phpr)3)2]n, multiples of 1490 g mol-1).