Evidence for a molten globule state in an oligomeric plant protein

Three different mustard globulin (isoforms), purified and isolated, using three extraction solutions (differing in ionic strength) were studied for structural integrity as a function of pH. Evidence of a molten globule state (a reversible intermediary state between the native and fully denatured forms) was obtained. This phenomenon may ultimately prove to be important in the translocation of these proteins across biological membranes at the time of their biosynthesis. Circular dichroism, hydrophobic probe, fluorescence spectral scans and differential scanning calorimetry were used to study this phenomenon. Secondary and tertiary structures (circular dichroism (CD) data) were found to be similar for globulins from higher ionic strength extractions, but different from the globulin from distilled water extraction; however, for all three isoforms, little change in secondary structure fractions as a function of pH was observed. Changes in tertiary structure (near-UV CD and intrinsic fluorescence data) as a function of pH were observed for all three globulin isoforms with greatest changes in tertiary structure being seen in the acidic pH range, i.e. 3–5. In contrast, all globulins were shown to undergo the least conformational change in the pH range of 6–9.