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

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• 作者：Masako Nagai ; Shigenori Nagatomo ; Yukifumi Nagai ; Kenichi Ohkubo ; Kiyohiro Imai ; Teizo Kitagawa
• 刊名：Biochemistry
• 出版年：2012
• 出版时间：July 31, 2012
• 年：2012
• 卷：51
• 期：30
• 页码：5932-5941
• 全文大小：538K
• 年卷期：v.51,no.30(July 31, 2012)
• ISSN：1520-4995

文摘

The aromatic residues such as tryptophan (Trp) 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 鈫?T quaternary structure transition. In Hb A, there are three Trp residues per 伪尾 dimer: at 伪14, 尾15, and 尾37. To evaluate their individual contributions to the R 鈫?T spectral changes, we produced three mutant hemoglobins in E. coli; rHb (伪14Trp鈫扡eu), rHb (尾15Trp鈫扡eu), and rHb (尾37Trp鈫扝is). 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 Trp residues were extracted by the difference calculations between Hb A and the mutants. 伪14 and 尾15Trp exhibited negative CD bands in both oxy- and deoxy-Hb A, whereas 尾37Trp showed positive CD bands in oxy-Hb A but decreased intensity in deoxy-form. These differences in CD spectra among the three Trp residues in Hb A were ascribed to surrounding hydrophobicity by examining the spectral changes of a model compound of Trp, N-acetyl-l-Trp ethyl ester, in various solvents. Intensity enhancement of Trp UVRR bands upon the R 鈫?T transition was ascribed mostly to the hydrogen-bond formation of 尾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.