Side-chain assignments of methyl-containing residues in a uniformly 13C-labeled hemoglobin in the carbonmonoxy form

Sequence-specific assignment of the methyl groups in large proteins can be obtained from an MQ-(H)CC(m)H(m)-TOCSY experiment on uniformly (13)C-labeled proteins without deuteration (Yang et al., 2004). Here the procedure is further demonstrated on a uniformly (13)C-labeled alpha-chain or beta-chain of human normal adult hemoglobin (65 kDa) in the carbonmonoxy form. In addition, a strategy is presented for assigning protons of methyl-containing residues of uniformly (13)C-labeled large proteins, on the basis of prior methyl assignments based on MQ-(H)CCH-TOCSY and H(C)C(m)H(m)-TOCSY experiments. Assignment of about 80% of the side-chain resonances of methyl-containing residues of carbonmonoxyhemoglobin has been obtained.