Energetics of the Thermal Transitions of RNA 1 and RNA 4 of Alfalfa‐Mosaic Virus in the Presence and Absence of Coat Protein

Abstract

Transition enthalpies of one genomic RNA, RNA 1. and the subgenomic RNA 4 of alfalfa mosaic virus were determined by scanning microcalorimetry in the absence and presence of various amounts of coat protein in a buffer containing 10 mM NaH2PO4, 1 mM EDTA, 1mM NaH3, pH 7.0. Analogous measurements were performed with the top and bottom components of alfalfa mosaic virus in the same buffer and with the coat protein in water. A comparison of the various enthalpies showed that the energetics of the protein–nucleic‐acid interaction are different for RNA 1 and RNA 4. The thermally induced transition of the coat protein in water is characterized by an enthalpy of 418 ± 67 kJ (100 ± 16 kcal) mol−1 at 38.9°C and a heat capacity change of approximately 6.7 kJ (1.6 kcal) mol−1 K−1. The unfolding pattern of the native bottom component of alfalfa mosaic virus, which consists of RNA 1 encapsulated by 240 coat protein molecules, is similar to that of the protein alone in exhibiting a single sharp heat capacity maximum and a heat capacity increase after the transition. The corresponding transition enthalpy is 523 ± 42 kJ (125 ± 10 kcal) · (mol protein)−1, the heat capacity change amounts to approximately 8 ± 1.7 kJ (1.9 ± 0.4 kcal) · (mol protein)−1, · K−1. An analogous transition curve results from unfolding of native top component fragments of alfalfa mosaic virus, which comprise two RNA 4 molecules per 132 protein molecules. The enthalpies involved in the unfolding process are 489 ± 42 kJ (117 ± 10 kcal) · (mol protein)−1, the heat capacity change 9.6 ± 2.5 kJ (2.3 + 0.6 kcal) · (mol protein)−1· K−1. The noticeable differences to the transition of the pure protein are the transition temperatures of 49.5°C and 47.9°C for the bottom and top components, respectively. The transition enthalpies of all protein · nucleic‐acid complexes are smaller than the sum of the transition enthalpies of the single components. This finding has been used to estimate enthalpies of interaction per mole of protein. The characteristic feature of these values is the fact that they all are positive, which indicates a decrease of the internal energy of the system on protein–nucleic‐acid interaction.