Hydrogen exchange in the carbon monoxide complex of soybean leghemoglobin.
Hydrogen/deuterium exchange rates for individual amide protons have been measured for the carbon monoxide complex of soybean leghemoglobin. Fast two-dimensional NOESY experiments were performed, with 5.2-min data-collection time for each spectrum, which made possible the measurement of NOE cross-peaks of relatively rapidly exchanging amide protons at early time points. Exchange rates were measured for 61 backbone amides, the protection factors were calculated to provide information on the packing and local stability of the protein. The data are consistent with the presence of transient cooperative local unfolding of helical segments. The B-, E-, G- and H-helices have extensive regions of slow-, medium- and fast-exchanging amide protons. For each of these helices, there is a progressive decrease in protection on moving from the helix center to the termini. This is consistent with a stable helix center, with dynamic fraying at the ends. Amide exchange from the A-helix and C-helix is rapid except in small local regions. The F-helix, which is located on the proximal side of the heme pocket and is well formed in solution as demonstrated by characteristic medium range NOE connectivities [Morikis, D. Lepre, C.A. & Wright, P.E. (1994) Eur. J. Biochem. 219, 611-626], exhibits fast exchange for all amide protons. The implied flexibility and low stability of the F-helix may be functionally important in facilitating movement of the helix upon ligand binding. Fast exchange has also been observed for all amide protons in the CE-loop and in turns, as expected for flexible or solvent exposed regions. A strong tertiary contact has been established between the A-, G- and H-helices by the presence of a slowly exchanging indole N epsilon H of Trp129.
Morikis
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Department of Molecular Biology, Scripps Research Institute, La Jolla, CA 92037, USA.
Wright
P E
PE
D000595:Amino Acid Sequence; D002248:Carbon Monoxide / Q000737:chemistry*; D003903:Deuterium / Q000737:chemistry; D007874:Leghemoglobin / Q000737:chemistry*; D008969:Molecular Sequence Data; D013025:Glycine max / Q000737:chemistry*
D007874:Leghemoglobin; D002248:Carbon Monoxide; D003903:Deuterium