Bhuta A, Quiggle K, Ott T, Ringer D, Chládek S
Biochemistry. 1981 Jan 6;20(1):8-15. doi: 10.1021/bi00504a002.
The substrate specificity of the acceptor site of peptidyltransferase of Escherichia coli 70S ribosomes was investigated in the fMet-tRNA.A-U-G.70S ribosome and AcPhe-tRNA.poly(U).70S ribosome systems by using a series of 2'- and 3'-aminoacyldinucleoside phosphates as acceptors. These chemically synthesized compounds are analogues of the 3' termini of either 2'(3')-, 2'-, or 3'-aminoacyl transfer ribonucleic acids (AA-tRNAs) of the types C-A-aa, C-2'-dA-aa, C-3'-dA-aa, C-3'-dA-3'-NH-aa, and C-2'-dA-2'-NH-aa (aa = Phe, D-Phe, Lys, Leu, Ala, Glu, Pro, Gly, Asp, Met, and alpha-aminoisobutyryl). It was found that the 3'-aminoacyl derivatives of optically active amino acids are much better acceptors of N-formyl-L-methionine (fMet) or N-acetyl-L-phenylalanine (AcPhe) residues than the isomeric 2'-aminoacyl derivatives with affinity constant ratios (KM 2'/3') greater than 100. Likewise, C-A(D-Phe) is a weaker acceptor than the corresponding L derivative C-A-Phe. In contrast, all glycyl derivatives (C-2'-dA-Gly, C-3'-dA-Gly, C-3'-dA-3'-NH-Gly and C-2'-dA-2'-NH-Gly) are good acceptors of the fMet residue, with ratios (KM 2'/3') of approximately 2. On the basis of these results, a model for the stereochemical control of the peptidyl-transferase reaction is proposed. It assigns a major role to the orientation of the amino acid side chain in 2'- or 3'-AA-tRNA. A detailed model of the interaction of the acceptor terminus of 3'-AA-tRNA with the acceptor site of peptidyltransferase is also proposed. The model is strikingly similar to those for the active sites of proteolytic enzymes.
在甲硫氨酰 - tRNA.A - U - G.70S核糖体和乙酰苯丙氨酰 - tRNA.poly(U).70S核糖体系统中，通过使用一系列2'-和3'-氨基酰二核苷磷酸作为受体，研究了大肠杆菌70S核糖体肽基转移酶受体位点的底物特异性。这些化学合成的化合物是C - A - aa、C - 2'-dA - aa、C - 3'-dA - aa、C - 3'-dA - 3'-NH - aa和C - 2'-dA - 2'-NH - aa（aa = 苯丙氨酸、D - 苯丙氨酸、赖氨酸、亮氨酸、丙氨酸、谷氨酸、脯氨酸、甘氨酸、天冬氨酸、甲硫氨酸和α - 氨基异丁酰基）类型的2'(3') - 、2'-或3'-氨基酰转移核糖核酸（AA - tRNA）3'末端的类似物。结果发现，旋光性氨基酸的3'-氨基酰衍生物比具有大于100的亲和常数比（KM 2'/3'）的异构2'-氨基酰衍生物是更好的N - 甲酰 - L - 甲硫氨酸（fMet）或N - 乙酰 - L - 苯丙氨酸（AcPhe）残基的受体。同样，C - A(D - 苯丙氨酸)是比相应的L衍生物C - A - 苯丙氨酸更弱的受体。相比之下，所有甘氨酰衍生物（C - 2'-dA - 甘氨酸、C - 3'-dA - 甘氨酸、C - 3'-dA - 3'-NH - 甘氨酸和C - 2'-dA - 2'-NH - 甘氨酸）都是fMet残基的良好受体，其比值（KM 2'/3'）约为2。基于这些结果，提出了肽基转移酶反应立体化学控制的模型。它赋予了2'-或3'-AA - tRNA中氨基酸侧链取向主要作用。还提出了3'-AA - tRNA受体末端与肽基转移酶受体位点相互作用的详细模型。该模型与蛋白水解酶活性位点的模型惊人地相似。
