Venkatesan S, Moss B
Proc Natl Acad Sci U S A. 1982 Jan;79(2):340-4. doi: 10.1073/pnas.79.2.340.
Incubation of HeLa cell mRNA guanylyltransferase (GTP:mRNA guanylyltransferase, EC 2.7.7.50) with [alpha-32P]GTP and a divalent cation in the absence of an RNA acceptor results in the formation of a covalent enzyme-guanylate complex. The complex, after purification by phosphocellulose chromatography, can transfer its bound GMP moiety to pyrophosphate, regenerating GTP, or to the 5'-diphosphate end of poly(A), forming a cap structure G(5')pppA(pA)n. The GMP-polypeptide has a molecular weight of 65,000 and is stable to heating in the presence of sodium dodecyl sulfate. On the basis of the alkali-stable and acid-labile nature of the bond and its susceptibility to nucleophilic attack by hydroxylamine at low pH, the GMP-polypeptide linkage appears to be a phosphoamine bond. After digestion with trypsin, a single GMP-peptide was resolved by two dimensional electrophoresis and chromatography.
在没有RNA受体的情况下，将HeLa细胞mRNA鸟苷酸转移酶（GTP：mRNA鸟苷酸转移酶，EC 2.7.7.50）与[α-32P]GTP和二价阳离子一起温育，会导致形成共价酶-鸟苷酸复合物。该复合物经磷酸纤维素色谱纯化后，可将其结合的GMP部分转移至焦磷酸，再生GTP，或转移至聚（A）的5'-二磷酸末端，形成帽结构G(5')pppA(pA)n。GMP-多肽的分子量为65,000，在十二烷基硫酸钠存在下对加热稳定。基于该键对碱稳定而对酸不稳定的性质以及其在低pH下对羟胺亲核攻击的敏感性，GMP-多肽键似乎是一种磷胺键。用胰蛋白酶消化后，通过二维电泳和色谱法分离出单一的GMP-肽。
