Purification and Partial Characterization of Genetically Engineered Thiol Protease Over-expressed in Escherichia Coli and Analysis of Priming Effects on Free Amino Acid Accumulation in Pinus Taeoa Seeds
Abstract
The overall objective of an ongoing project is to determine the physiological and genetic mechanisms responsible for the invigoration of loblolly pine (Pinus taeda L.) seeds subjected to controlled water stress through solid matrix priming (SMP). It is hypothesized that the invigoration is due to a change in the sequence of events in the germinating seed where storage protein degradation and mobilization which normally commence after germination are stimulated to occur before radical emergence. The mobilization of protein reserves provides: osmotically active substances for osmotic adjustment leading to increased capacity of the embryo to grow and the components for synthesis of new proteins for the growing plant. It is also hypothesized that thiol protease (TP) is a major enzyme involved in the degradation of storage proteins when seeds are primed, as it is known to degrade reserve proteins, to be active during germination and to be up-regulated by water stress. As one effort toward the overall objective, the first chapter concerns TP antigen production to make anti-TP antibodies so that it becomes possible to further elucidate TP functions at the translation and post-translation levels cellularly and subcellularly. In addition, the over-expressed TP protein provides a way to efficiently purify the enzyme by immunoaffinity chromatography from loblolly pine. The second chapter, without direct relations to Chapter I but as another step toward the overall goal, concerns SMP effects on free amino acid accumulation and the relationships among free amino acid accumulation, TP activity increase and decrease of water potential.
Collections
- OSU Theses [15752]