Millisecond time scale conformational flexibility in a hyperthermophile protein at ambient temperature

Rubredoxin from the hyperthermophile Pyrococcus furiosus is the most thermo stable protein characterized to date with an estimated global unfolding rat e of 10(-6) s(-1) at 100 degrees C. In marked contrast to these slow global dynamics, hydrogen exchange experiments here demonstrate that conformation al opening for solvent access occurs in the approximate to millisecond time frame or faster at 28 degrees C for all amide positions. Under these condi tions all backbone amides with exchange protection factors between 10(4) an d 10(6), for which EX2 exchange kinetics were directly verified, have excha nge activation energy values within 2-3 kcal/mol of that observed for unstr uctured peptides. The conformational flexibility of this protein is thus su fficient for water and base catalyst access to the exchanging amide with qu ite limited structural disruption. The common hypothesis that enhanced conf ormational rigidity in the folded native state underlies the increased ther mal stability of hyperthermophile proteins is not supported by these data.