77K的时间分辨步进扫描FTIR差异光谱已用于研究集胞藻属的光系统I(PSI)。PCC6803具有四个高电位,1,4-萘醌(NQs)掺入A1结合位点。掺入的醌是2-氯-NQ(2ClNQ),2-溴-NQ(2BrNQ),2,3-二氯-NQ(Cl2NQ),和2,3-二溴-NQ(Br2NQ)。为了完整起见,还并入并研究了2-甲基-NQ(2MNQ)。以前,在中研究了掺入相同醌的PSI,所谓的,阴离子光谱区域介于1550和1400cm-1之间(Agarwala等人。“生物生物学报”1864(1):148918,2023)。在这里,我们专注于先前未探索的1400-1200cm-1光谱区域中的光谱。在该区域中,鉴定了几条带,并将其分配给掺入的醌的中性状态。这是重要的,因为在过去已经证明在规则的1700-1600cm-1区域中鉴定中性状态醌带是困难的。对于PSI中的中性PhQ,强烈的带出现在〜1300cm-1。对于对称的双取代NQs(Cl2NQ/Br2NQ),分别在〜1280/1269cm-1处发现了单个强烈的中性态带。对于两个单取代的NQ,2ClNQ和2BrNQ,然而,在〜1280和〜1250cm-1处分别观察到两个中性态条带。这些来自时间分辨光谱的观察结果与从THF中NQ的吸收光谱得出的结论非常吻合,这些也在这里介绍。进行基于密度泛函理论的振动频率计算,从而可以鉴定与中性状态醌带相关的正常模式。
Time-resolved step-scan FTIR difference spectroscopy at 77 K has been used to
study photosystem I (PSI) from Synechocystis sp. PCC 6803 with four high-potential, 1,4-naphthoquinones (NQs) incorporated into the A1 binding site. The incorporated quinones are 2-chloro-NQ (2ClNQ), 2-bromo-NQ (2BrNQ), 2,3-dichloro-NQ (Cl2NQ), and 2,3-dibromo-NQ (Br2NQ). For completeness 2-methyl-NQ (2MNQ) was also incorporated and studied. Previously, PSI with the same quinones incorporated were studied in the, so-called, anion spectral region between 1550 and 1400 cm-1 (Agarwala et al. in Biochim Biophys Acta 1864(1):148918, 2023). Here we focus on spectra in the previously unexplored 1400-1200 cm-1 spectral region. In this region several bands are identified and assigned to the neutral state of the incorporated quinones. This is important as identification of neutral state quinone bands in the regular 1700-1600 cm-1 region has proven difficult in the past. For neutral PhQ in PSI a broad, intense band appears at ~ 1300 cm-1. For the symmetric di-substituted NQs (Cl2NQ/Br2NQ) a single intense neutral state band is found at ~ 1280/1269 cm-1, respectively. For both mono-substituted NQs, 2ClNQ and 2BrNQ, however, two neutral state bands are observed at ~ 1280 and ~ 1250 cm-1, respectively. These observations from time-resolved spectra agree well with conclusions drawn from absorption spectra of the NQs in THF, which are also presented here. Density functional theory based vibrational frequency calculations were undertaken allowing an identification of the normal modes associated with the neutral state quinone bands.