Nonspecific hydrophobic interactions stabilize an equilibrium intermediate of apomyoglobin at a key position within the AGH region.
Acid-induced unfolding of apomyoglobin (apoMb) proceeds in a multistate process involving at least one equilibrium intermediate (I) at pH 4.2. The structure of the I form has been investigated thoroughly, with significant effort devoted to identifying potentially stabilizing native contacts. Here, we test whether rigid side-chain packing interactions like those in holomyoglobin persist at a buried position, Met-131, within the low-pH apoMb intermediate. We have measured the urea-induced unfolding transitions of overpacking, underpacking, and polar substitutions of Met-131 to determine the effect on the stability of the native and intermediate states of apoMb. Whereas underpacking substitutions should destabilize the I form irrespective of the degree of native side-chain-packing interactions, we anticipate that overpacking replacements might show opposite effects in a tightly packed environment, compared with a region lacking native side-chain packing interactions. We observe that, whereas underpacking and polar substitutions destabilize the I form, overpacking substitutions are stabilizing, implying that I is structurally plastic. We also report a strong correlation between the I state unfolding free energies and side-chain transfer free energies from water to octanol. Our results suggest that, whereas side-chain hydrophobicity is important for the stability of the I form, specific side-chain packing interactions are not.
T. C. Jenkins Department of Biophysics, The Johns Hopkins University, 3400 North Charles Street, Baltimore, MD 21218, USA.
Angela M
AM
Bertagna
Doug
D
Barrick
D000143:Acids / Q000737:chemistry; D001059:Apoproteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D057927:Hydrophobic and Hydrophilic Interactions; D009211:Myoglobin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D011489:Protein Denaturation; D017434:Protein Structure, Tertiary; D014508:Urea / Q000737:chemistry
D000143:Acids; D001059:Apoproteins; D009211:Myoglobin; C010857:apomyoglobin; D014508:Urea