pKa measurements from nuclear magnetic resonance of tyrosine-150 in class C beta-lactamase.
pKa measurements from nuclear magnetic resonance of tyrosine-150 in class C beta-lactamase.
复制标题
C 类 β-内酰胺酶中 150 酪氨酸的核磁共振 pKa 测量值。
DOI：
10.1042/bj20021447
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发表时间：
2003
期刊：
影响因子：
--
通讯作者：
M. Ishiguro
中科院分区：
文献类型：
--
作者：
Y. Kato;T. Iwashita;K. Masuda;Y. Oyama;M. Ishiguro
用~(13)C-核磁共振波谱测定了野生型和突变C类β-内酰胺酶中Tyr(150)(Y150)残基的pKa值。用(13)C标记的L酪氨酸([苯酚-4-(13)C]酪氨酸)取代野生型和突变型内酰胺酶的酪氨酸残基，以选择性地观察酪氨酸残基。将野生型和K67C突变(Lys(67)-&Gt；Cys)酶的光谱与Y150C突变的内酰胺酶光谱进行比较，以鉴定来自Tyr(150)的信号。滴定实验表明，野生型酶中Tyr(150)共振的化学位移在0.1p.m的范围内几乎不变。结果表明，该残基在无底物状态下的pK(A)远大于11。根据对X-射线衍射物结构的溶剂可及性计算，TYR(150)的酚氧位于Lys(315)和Lys(67)氨基附近，似乎具有较低的溶剂可及性。这些结果表明，在天然酶中，弗氏柠檬酸杆菌GN346 C类β-内酰胺酶中的Tyr(150)被质子化，当Tyr(150)失去一个质子时，一个来自Lys(67)的质子将取代它。因此，在整个滴定过程中，Tyr(150)将被质子化。
13C-NMR spectroscopy was used to estimate the p K a values for the Tyr(150) (Y150) residue in wild-type and mutant class C beta-lactamases. The tyrosine residues of the wild-type and mutant lactamases were replaced with (13)C-labelled L-tyrosine ([ phenol -4-(13)C]tyrosine) in order to observe the tyrosine residues selectively. Spectra of the wild-type and K67C mutant (Lys(67)-->Cys) enzyme were compared with the Y150C mutant lactamase spectra to identify the signal originating from Tyr(150). Titration experiments showed that the chemical shift of the Tyr(150) resonance in the wild-type enzyme is almost invariant in a range of 0.1 p.p.m. up to pH 11 and showed that the p K (a) of this residue is well above 11 in the substrate-free form. According to solvent accessibility calculations on X-ray-derived structures, the phenolic oxygen of Tyr(150), which is near the amino groups of Lys(315) and Lys(67), appears to have low solvent accessibility. These results suggest that, in the native enzyme, Tyr(150) in class C beta-lactamase of Citrobacter freundii GN346 is protonated and that when Tyr(150) loses a proton, a proton from Lys(67) would replace it. Consequently, Tyr(150) would be protonated during the entire titration.
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影响因子：
2.9
作者：
LOBKOVSKY, E;BILLINGS, EM;KNOX, JR
通讯作者：
KNOX, JR
影响因子：
2.9
作者：
Dyson, HJ;Jeng, MF;Holmgren, A
通讯作者：
Holmgren, A
β-二级和溶剂氘动力学同位素对链霉菌 R61 DD-肽酶催化作用的影响：与结构相似的 C 类 β-内酰胺酶的比较。
DOI：
10.1021/bi982308x
发表时间：
1999
期刊：
Biochemistry.
影响因子：
--
作者：
Adediran,SA;Pratt,RF
通讯作者：
Pratt,RF
影响因子：
--
作者：
Caselli, E;Powers, RA;Shoichet, BK
通讯作者：
Shoichet, BK