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Human cytoplasmic serine hydroxymethyltransferase is an mRNA binding protein

机译：人胞质丝氨酸羟甲基转移酶是一种mRNA结合蛋白

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摘要

The 5' untranslated region (UTR) of the human cytoplasmic serine hydroxymethyltransferase (cSHMT) message is alternatively spliced, creating a full-length 5' UTR (LUTR) encoded within exons 1-3 and a shorter UTR (SUTR) that results from excision of exon 2. The role of the 5' UTRs in cSHMT expression was investigated by fusing the cSHMT 5' UTRs to the 5' end of the luciferase gene. Human cSHMT protein at 10 mu M inhibits in vitro translation of cSHMT 5' UTR-luciferase fusion mRNA templates by more than 90%, but does not inhibit translation of the luciferase message lacking the UTR. Translation inhibition is independent of amino acid and folate substrate binding to the cSHMT enzyme. The cSHMT SUTR-luciferase mRNA binds to the cSHMT.glycine.5-formyltetrahydrofolate ternary complex with an apparent K-d of 10 mu M. Gel mobility shift assays demonstrate that the human cSHMT protein binds to the cSHMT LUTR-luciferase fusion mRNA in the presence and absence of glycine and 5-formyltetrahydrofolate pentaglutamate. The fusion cSHMT SUTR-luciferase message at 65 mu M inhibits the cSHMT-calalyzed cleavage of allothreonine as a partial mixed type inhibitor, reducing both k(cat) and K-m by 40 and 75%, respectively, while tRNA has no effect on cSHMT catalysis. These studies indicate that the cSHMT protein can bind mRNA, and displays increased affinity for the 5' untranslated region of its mRNA. [References: 32]
机译：交替剪接人胞质丝氨酸羟甲基转移酶（cSHMT）信息的5'非翻译区（UTR），创建在外显子1-3内编码的全长5'UTR（LUTR）和因切除而产生的较短UTR（SUTR）通过将cSHMT 5'UTR融合到萤光素酶基因的5'末端，研究了5'UTR在cSHMT表达中的作用。 10μM处的人cSHMT蛋白抑制cSHMT 5'UTR荧光素酶融合mRNA模板的体外翻译超过90％，但不抑制缺少UTR的荧光素酶信息的翻译。翻译抑制独立于氨基酸和叶酸底物与cSHMT酶的结合。 cSHMT SUTR-萤光素酶mRNA与cSHMT.glycine.5-甲酰基四氢叶酸三元复合物结合，表观Kd为10μM。凝胶迁移率变动分析表明，在存在和存在下，人cSHMT蛋白与cSHMT LUTR-萤光素酶融合mRNA结合。没有甘氨酸和5-甲酰基四氢叶酸五谷氨酸。融合cSHMT SUTR-荧光素酶信息在65μM时抑制了cSHMT催化的作为部分混合型抑制剂的Allallreonine裂解，使k（cat）和Km分别降低了40％和75％，而tRNA对cSHMT催化没有影响。这些研究表明，cSHMT蛋白可以结合mRNA，并且对其mRNA 5'非翻译区的亲和力增强。 [参考：32]

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