Megan M. Aoki , Anna B. Kisiala , Scott C. Farrow , Craig R. Brunetti , Robert J. Huber , R.J. Neil Emery
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Thus, we performed bioinformatic analyses and a biochemical characterization of a LOG ortholog from <em>Dictyostelium discoideum</em>, a soil-dwelling amoeba, which produces CKs during unicellular growth and multicellular development. We show that <em>Dd</em>Log exhibits LOG/PRH activity on two CK-NTs, <em>N</em><sup><em>6</em></sup>-isopentenyladenosine-5′-monophosphate (iPMP) and <em>N</em><sup><em>6</em></sup>-benzyladenosine-5′-monophosphate (BAMP), and on adenosine 5′-monophosphate (AMP) but not on 3′, 5′-cyclic adenosine-monophosphate (cAMP). Additionally, there were higher turnover rates for CK-NTs over AMP. 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引用次数: 0
摘要
孤独者(LOG)蛋白是一种磷酸核糖水解酶(PRHs),是植物和非植物细胞分裂素(CK)产生生物体中的关键细胞分裂素(CK)活化酶。在 CK 生物合成过程中,LOG 催化前体 CK 核苷酸(CK-NTs)转化为具有生物活性的游离碱基形式。在细菌、古菌、藻类和真菌中都检测到了 LOG/PRH 活性。然而,在这些生物体中,CK-NT 和非 CK-NT(如腺嘌呤-NT)的 LOG/PRH 活性尚未同时得到评估,因此对 LOG 的底物特异性了解有限。因此,我们对来自盘状竹荪的 LOG 直向同源物进行了生物信息学分析和生化鉴定,盘状竹荪是一种生活在土壤中的变形虫,在单细胞生长和多细胞发育过程中会产生 CKs。我们发现,DdLog 对两种 CK-NTs(N6-异戊烯基腺苷-5′-单磷酸(iPMP)和 N6-苄基腺苷-5′-单磷酸(BAMP))以及腺苷-5′-单磷酸(AMP)具有 LOG/PRH 活性,但对 3′,5′-环腺苷-单磷酸(cAMP)不具有 LOG/PRH 活性。此外,CK-NTs 的周转率高于 AMP。这些发现共同证实了 DdLog 起着 CK 激活酶的作用;然而,与植物 LOG 不同的是,它对其他底物(如 AMP)保持着更广泛的特异性,这反映出它即使在多样化为 CK 激活酶后,仍保持着原来与 CK 无关的作用。
Biochemical characterization of a unique cytokinin and nucleotide phosphoribohydrolase Lonely Guy protein from Dictyostelium discoideum
Lonely guy (LOG) proteins are phosphoribohydrolases (PRHs) that are key cytokinin (CK)-activating enzymes in plant and non-plant CK-producing organisms. During CK biosynthesis, LOGs catalyze the conversion of precursor CK-nucleotides (CK-NTs) to biologically active free base forms. LOG/PRH activity has been detected in bacteria, archaea, algae, and fungi. However, in these organisms, the LOG/PRH activity for CK-NTs and non-CK-NTs (e.g., adenine-NTs) has not been assessed simultaneously, which leaves limited knowledge about the substrate specificity of LOGs. Thus, we performed bioinformatic analyses and a biochemical characterization of a LOG ortholog from Dictyostelium discoideum, a soil-dwelling amoeba, which produces CKs during unicellular growth and multicellular development. We show that DdLog exhibits LOG/PRH activity on two CK-NTs, N6-isopentenyladenosine-5′-monophosphate (iPMP) and N6-benzyladenosine-5′-monophosphate (BAMP), and on adenosine 5′-monophosphate (AMP) but not on 3′, 5′-cyclic adenosine-monophosphate (cAMP). Additionally, there were higher turnover rates for CK-NTs over AMP. Together, these findings confirm that DdLog acts as a CK-activating enzyme; however, in contrast to plant LOGs, it maintains a wider specificity for other substrates (e.g., AMP) reflecting it has maintained its original, non-CK related role even after diversifying into a CK-activating enzyme.
期刊介绍:
Open access, online only, peer-reviewed international journal in the Life Sciences, established in 2014 Biochemistry and Biophysics Reports (BB Reports) publishes original research in all aspects of Biochemistry, Biophysics and related areas like Molecular and Cell Biology. BB Reports welcomes solid though more preliminary, descriptive and small scale results if they have the potential to stimulate and/or contribute to future research, leading to new insights or hypothesis. Primary criteria for acceptance is that the work is original, scientifically and technically sound and provides valuable knowledge to life sciences research. We strongly believe all results deserve to be published and documented for the advancement of science. BB Reports specifically appreciates receiving reports on: Negative results, Replication studies, Reanalysis of previous datasets.