Circular-dichroism and absorption spectroscopic studies on specific aromatic residues involved in the different modes of aggregation of tobacco-mosaic-virus protein.

Conformational changes accompanying the different modes of aggregation of tobacco mosaic virus protein (TMV-protein) were investigated using circular dichroism (CD) and absorption difference spectra in the range of aromatic absorption. Comparing wild-type protein and mutant Ni 2068 (Tyr-139 leads to Cys-139) a tentative localization of aromatic amino acids in the three-dimensional structure is rendered possible. In all modes of aggregation the CD spectra are determined by intrasubunit interactions between aromatic residues, in particular Trp-17 and Trp-52 as well as Tyr-70, Tyr-72 and Tyr-139. The Trp-17-Trp-52 interaction was found to be highly sensitive towards changes of the quaternary structure especially with respect to helical aggregates. This suggests that the environment of the two tryptophan residues is of crucial importance in the three-dimensional structure of the subunit; in the course of aggregation intersubunit interactions compete with the specific intrasubunit Trp-17--Trp52 interactions. It is suggested that Try-70 and Tyr-72 form hydrogen bonds in a strongly hydrophobic environment. Formation of the double disc decreases the rotatory strength, pointing to an increase in conformational flexibility. Spectroscopic and chemical evidence prove that Tyr-70, Tyr-72 and Tyr-139 are in close neighbourhood. Double disc formation by lowering the pH (pH 8 LEADS TO 6.9, I = 0.1 M) or increasing the ionic strength (pH 8, I = 0.1 LEADS TO 0.6 M) is reflected by identical spectral effects in the environment of Tyr-70 - Tyr-72. However the interaction between Trp-17 and Trp-52 indicates significant differences in the conformation which may be important for the formation of higher aggregates, i.e. 'lockwashers', helices, and 'stacked discs'.