Barry T. Nall

Molecular mechanisms governing reading frame choice of immunoglobulin diversity genes

Abstract The key elements of antigen-binding immunoglobulin (lg) variable regions are primarily encoded by diversity (D H ) genes. Although D H genes can be used in all three reading frames, the majority of peripheral lg molecules carry a D H element in a single reading frame. Here, Frank Raaphorst, C. Raman, Barry Nall and Judy Teale discuss the particular demands imposed by the structure of the antigen-binding site, which determine the choice of reading frame.

Crystallization of yeast iso-2-cytochrome c using a novel hair seeding technique☆

A hair seeding technique has been developed to obtain diffraction quality crystals of yeast (Saccharomyces cerevisiae) iso-2-cytochrome c, a model for studies of protein folding and biological electron transfer reactions. Deep red crystals of this protein were obtained from 88 to 92% saturated solutions of ammonium sulfate containing 20 mg protein/ml, 0.1 M-sodium phoshate, 0.3 M-sodium chloride, 0.04 M-dithiothreitol and adjusted to phosphate, 0.3 M-sodium chloride, 0.04 M-dithiothreitol and adjusted to pH 6.0. Rapid crystal growth was observed, but only along the path of the seeding hair stroke. The space group is P4(3)2(1)2 (or P4(1)2(1)2) with a = b = 36.4 A, c = 137.8 A (1 A = 0.1 nm) and Z = 8. Crystals are stable in the X-ray beam for more than 10 days and diffract to at least 2.5 A resolution. The same hair seeding methodology has proven useful in obtaining crystals of specifically designed mutant iso-2 proteins and in other protein systems where consistent crystal growth had previously proven difficult to attain.

Proline Isomerization and Protein Folding

A reaction as complex as folding of a protein must involve a variety specific chemical processes. Hydrogen bonds and salt bridges are formed, and perhaps broken and interchanged. Solvent-induced hydrophobic associations may occur, or even a general hydrophobic collapse of the polypeptide to a less than fully ordered “globule”. Polypeptide chains might become entangled with each other or with other chains and have to extricate themselves prior to further folding. Intricate shapes and structures may be constructed only to partially or fully unravel on transformation into other more stable forms. The objective of much experimental work on the process of folding has been to obtain direct evidence for some of these expectations. This has been a major challenge since many aspects of folding, rather than having the expected complexity, are, instead, well described by the simplest of chemical mechanisms: a two state process.