The acid-stable potato tuber proteins: Identification and isolation of invertase inhibitor and acid-stable potato hemagglutinins by high-performance liquid chromatography and isoelectrofocusing polyacrylamide gel electrophoresis

The objective of this research was to purify an inhibitor protein that binds to and reduces the activity of potato (Solanum tuberosum L.) tuber acid invertase. The investigation led to the following conclusions: (1) A single protein band on an isoelectrofocusing polyacrylamide gel (IEF-PAGE) was identified as the invertase inhibitor. The inhibitor was a protein with a molecular weight of approximately 17-18 kD and a pI of 5.1. (2) An acid-stable hemagglutinin was found to co-purify with the potato invertase inhibitor after elution from size exclusion, hydroxylapatite, and anion exchange chromatographies. Hemagglutinin activity was associated with a single band which had a pI of 5.6 on IEF-PAGE gels. A second hemagglutinin, which showed no affinity for either hydroxylapatite or DEAE-Sephadex, was isolated from the same preparation. The second hemagglutinin was associated with a protein with a pI of 8.4. Both hemagglutinins have a molecular weight of approximately 17-18 kD, as does invertase inhibitor. (3) The three proteins co-purified along with other proteins of similar molecular weight, but different pI values. The presence of multiple proteins accompanying the invertase inhibitor was obscured on discontinuous polyacrylamide gels (SDS-PAGE). (4) The proteins were separated by nonideal size exclusion chromatography (nSEC). The process is different from that of normal size exclusion chromatography in that the proteins were sorted on the basis of pI in addition to molecular weight. (5) Potato tuber invertase was characterized as a multimeric protein with a molecular weight of approximately 240 kD and a monomer weight of 38 kD.