Evaluating protein prenylation of human and viral CaaX sequences using a humanized yeast system.

IF 4 3区 医学 Q2 CELL BIOLOGY Disease Models & Mechanisms Pub Date : 2024-05-01 Epub Date: 2024-05-31 DOI:10.1242/dmm.050516
Emily R Hildebrandt, Anushka Sarkar, Rajani Ravishankar, June H Kim, Walter K Schmidt
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Abstract

Prenylated proteins are prevalent in eukaryotic biology (∼1-2% of proteins) and are associated with human disease, including cancer, premature aging and infections. Prenylated proteins with a C-terminal CaaX sequence are targeted by CaaX-type prenyltransferases and proteases. To aid investigations of these enzymes and their targets, we developed Saccharomyces cerevisiae strains that express these human enzymes instead of their yeast counterparts. These strains were developed in part to explore human prenyltransferase specificity because of findings that yeast FTase has expanded specificity for sequences deviating from the CaaX consensus (i.e. atypical sequence and length). The humanized yeast strains displayed robust prenyltransferase activity against CaaX sequences derived from human and pathogen proteins containing typical and atypical CaaX sequences. The system also recapitulated prenylation of heterologously expressed human proteins (i.e. HRas and DNAJA2). These results reveal that substrate specificity is conserved for yeast and human farnesyltransferases but is less conserved for type I geranylgeranyltransferases. These yeast systems can be easily adapted for investigating the prenylomes of other organisms and are valuable new tools for helping define the human prenylome, which includes physiologically important proteins for which the CaaX modification status is unknown.

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利用人源化酵母系统评估人类和病毒 CaaX 序列的蛋白质前酰化。
在真核生物中,前炔基化蛋白质非常普遍(占蛋白质的 1%~2%),并且与人类疾病(包括癌症、早衰和感染)有关。具有 C 端 CaaX 序列的前酰化蛋白质是 CaaX 型前酰转移酶和蛋白酶的靶标。为了帮助研究这些酶及其靶标,我们开发了表达这些人类酶而不是酵母对应酶的酿酒酵母菌株。开发这些菌株的部分目的是为了探索人类前酰转移酶的特异性,因为我们发现酵母 FTase 对偏离 CaaX 共识(即非典型序列和长度)的序列具有更强的特异性。人源化酵母菌株对来自人类和病原体蛋白质的含有典型和非典型 CaaX 序列的 CaaX 序列具有很强的前基转移酶活性。该系统还重现了异源表达的人类蛋白质(即 HRas 和 DNAJA2)的前酰化。这些结果表明,酵母和人类法尼基转移酶的底物特异性是一致的,而 I 型香叶基转移酶的底物特异性则不太一致。这些酵母系统可以很容易地用于研究其他生物的前酰体,而且是帮助确定人类前酰体的宝贵新工具,人类前酰体包括CaaX修饰状态未知的重要生理蛋白质。
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来源期刊
Disease Models & Mechanisms
Disease Models & Mechanisms 医学-病理学
CiteScore
6.60
自引率
7.00%
发文量
203
审稿时长
6-12 weeks
期刊介绍: Disease Models & Mechanisms (DMM) is an online Open Access journal focusing on the use of model systems to better understand, diagnose and treat human disease.
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