Novel tyrosine markers in Raman spectra of wild-type and mutant (Y21M and Y24M) Ff virions indicate unusual environments for coat protein phenoxyls.
Novel tyrosine markers in Raman spectra of wild-type and mutant (Y21M and Y24M) Ff virions indicate unusual environments for coat protein phenoxyls.
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DOI：
10.1021/bi00171a001
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发表时间：
1994-02
期刊：
影响因子：
2.9
通讯作者：
S. Overman;K. L. Aubrey;N. Vispo;G. Cesareni;G. Thomas
中科院分区：
文献类型：
--
作者：
S. Overman;K. L. Aubrey;N. Vispo;G. Cesareni;G. Thomas
酪氨酸侧链产生一对独特的拉曼谱带--在850和830 cm-1附近的费米双峰--其相对强度可诊断酚受体和供体原子的氢键状态[Siamwiza等（1975）Biochemistry 14，4870-4876]。这种结构相关性已被广泛测试，并被广泛用作球状蛋白及其组装体中酪氨酸相互作用的指标。然而，在Ff丝状病毒（fd，f1，M13）的表观费米双峰强度比（I853/I826约4.0）是远远大于最大预测或观察到的其他蛋白质。为了理解这一反常现象，我们重新评估了Ff中费米二重态分配的基础。我们报告的拉曼光谱的特定位点突变体的Ff，其中一个（Y21 M和Y24 M）或两个（Y21 F/Y24 S）酪氨酸的外壳蛋白亚基（pVIII）已发生突变。这些拉曼数据，以及从携带pVIII中的残基特异性酪氨酰（Y-d4）和苯丙氨酰（F-d5）氘代的Ff病毒体获得的数据，最终证明Ff的853和826 cm-1谱带不构成典型的酪氨酸费米双峰：观察到的Ff的826 cm-1拉曼谱带不是由于pVIII的酪氨酸，而是由于pVIII的苯丙氨酸残基。因此，853 cm-1拉曼谱带构成了蛋白质的拉曼光谱中“酪氨酸单线态”的第一个已知实例。这一发现的影响Ff病毒粒子的结构和其他蛋白质中的酪氨酸标志物的相关性进行了讨论。
The tyrosine side chain generates a pair of distinctive Raman bands--a Fermi doublet near 850 and 830 cm-1--with relative intensities diagnostic of hydrogen bonding states of the phenolic acceptor and donor atoms [Siamwiza et al. (1975) Biochemistry 14, 4870-4876]. This structural correlation has been tested extensively and is used widely as an indicator of tyrosine interactions in globular proteins and their assemblies. However, in Ff filamentous viruses (fd, f1, M13) the apparent Fermi doublet intensity ratio (I853/I826 approximately 4.0) is much greater than the maximum predicted or observed in other proteins. To understand this anomaly, we have reevaluated the basis for the Fermi doublet assignment in Ff. We report Raman spectra of site-specific mutants of Ff in which either one (Y21M and Y24M) or both (Y21F/Y24S) tyrosines of the coat protein subunit (pVIII) have been mutated. These Raman data, together with those obtained from Ff virions carrying residue-specific tyrosyl (Y-d4) and phenylalanyl (F-d5) deuterations in pVIII, demonstrate conclusively that the 853 and 826 cm-1 bands of Ff do not constitute a typical tyrosine Fermi doublet: The observed 826 cm-1 Raman band of Ff is due not to tyrosine but to phenylalanine residues of pVIII. The 853 cm-1 Raman band thus constitutes the first known example of a "tyrosine singlet" in the Raman spectrum of a protein. The implications of this finding for Ff virion structure and its relevance to tyrosine markers in other proteins are discussed.