The Mechanism of Folding of Dihydrofolate Reductase

点击翻译按钮获取中文摘要

9604678 Matthews The overall goal of this research is to understand the mechanism by which the amino acid sequence of a protein directs its rapid and efficient folding to its native conformation. A combination of biophysical and genetic engineering methods will be used to probe the mechanism of folding of homologous dihydrofolate reductases from a common bacterium, E. coli, and from humans. Circular dichroism, NMR, fluorescence and x-ray scattering studies will provide insight into the secondary and tertiary structures, as well as the dynamics and sizes of transient, partially-folded forms that have previously been observed to appear during the folding of both proteins. Ultrafast mixing will probe the collapse of unfolded forms and the development of nonpolar surfaces within 100 microseconds. Mutational analysis will probe the involvement of nonpolar side chains in several hydrophobic clusters that appear to play key roles in two classes of folding intermediates. Protein engineering will test the potential of fragments to fold. Finally, the role of a bacterial chaperone, GroEL, in influencing the folding of human dihydrofolate reductase will be investigated. Progress in understanding the structures and dynamics of partially folded species that appear during folding should provide insight into the process by which the amino acid sequence of a protein spontaneously directs the formation of its unique three dimensional structure. A solution to the protein folding problem would have an important effect on biology, biochemistry and the biotechnology industry. The folding and transport of extracellular proteins is often coordinated with their own synthesis or with the synthesis of other proteins with which they form complexes. Thus, folding plays a key role in the normal functioning of cells. Because the biological activities of proteins are directly related to their three dimensional structures, an understanding of the sequence/structure relationship is a critical component of the processing of the genetic information stored in the DNA sequence.