Near-UV Circular Dichroism and UV Resonance Raman Spectra of Individual Tryptophan Residues in Human Hemoglobin and Their Changes upon the Quaternary Structure Transition

作者：Nagai Masako^*; Nagatomo Shinori; Nagai Yukifitmi; Ohkubo Kenichi; Imai Kiyohiro; Kitagawa Teizo

来源：Biochemistry, 2012, 51(30): 5932-5941.

DOI：10.1021/bi300347x

The aromatic residues such as tryptophan (Tip) and tyrosine (Tyr) in human adult hemoglobin (Hb A) are known to contribute to near-UV circular dichroism (CD) and UV resonance Raman (RR) spectral changes upon the R T quaternary structure transition. In Hb A, there are three Tip residues per alpha beta dimer: at alpha 14, beta 15, and beta 37. To evaluate their individual contributions to the R -%26gt; T spectral changes, we produced three mutant hemoglobins in E. coli; rHb (alpha 14Trp -%26gt; Leu), rHb (beta 15Trp -%26gt; Leu), and rHb (beta 37Trp -%26gt; His). Near-UV CD and UVRR spectra of these mutant Hbs were compared with those of Hb A under solvent conditions where mutant rHbs exhibited significant cooperativity in oxygen binding. Near-UV CD and UVRR spectra for individual Tip residues were extracted by the difference calculations between Hb A and the mutants. alpha 14 and beta 15Trp exhibited negative CD bands in both oxy- and deoxy-Hb A, whereas beta 37Trp showed positive CD bands in oxy-Hb A but decreased intensity in deoxy-form. These differences in CD spectra among the three Tip residues in Hb A were ascribed to surrounding hydrophobicity by examining the spectral changes of a model compound of Tip, N-acetyl-L-Trp ethyl ester, in various solvents. Intensity enhancement of Trp UVRR bands upon the R -%26gt; T transition was ascribed mostly to the hydrogen-bond formation of beta 37Trp in deoxy-Hb A because similar UVRR spectral changes were detected with N-acetyl-L-Trp ethyl ester upon addition of a hydrogen-bond acceptor.

• 出版日期2012-7-31