Structural basis for the allosteric regulation of the SbtA bicarbonate transporter by the PII-like protein, SbtB, from Cyanobium sp. PCC7001
Structural basis for the allosteric regulation of the SbtA bicarbonate transporter by the PII-like protein, SbtB, from Cyanobium sp. PCC7001
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来自 Cyanobium sp. 的 PII 样蛋白 SbtB 对 SbtA 碳酸氢盐转运蛋白变构调节的结构基础。
DOI：
10.1101/762807
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发表时间：
2019
期刊：
影响因子：
--
通讯作者：
C. Jackson
中科院分区：
文献类型：
--
作者：
J. Kaczmarski;N. Hong;Bratati Mukherjee;Laura T. Wey;Loraine M. Rourke;Britta Förster;T. Peat;G. Price;C. Jackson
蓝藻已经进化出一套酶和无机碳（Ci）转运蛋白，其通过增加主要CO2固定酶Rubisco周围的CO2局部浓度来提高光合性能。这种CO2浓缩机制（CCM）是高度调节的，响应于光照/黑暗循环，并允许蓝藻在有限的Ci条件下茁壮成长。虽然CCM活性的转录控制是很好的理解，很少有人知道调节蛋白如何变构调节Ci转运蛋白响应不断变化的条件。蓝藻钠依赖性碳酸氢盐转运蛋白（SbtAs）被类PII调节蛋白（SbtB）抑制，抑制作用由腺苷酸调节。在这里，我们使用等温滴定量热法显示，SbtB从蓝藻属物种PCC 7001（SbtB 7001）结合AMP，ADP，cAMP和ATP与微摩尔范围的亲和力。载脂蛋白和核苷酸结合的SbtB 7001的X射线晶体结构显示，虽然AMP，ADP和cAMP对SbtB 7001结构的影响很小，但ATP的结合稳定了其他灵活的T环，并且灵活的C末端C环采用了几种不同的构象。我们还表明，ATP结合亲和力增加十倍，在Ca 2+的存在下，我们提出了一个X-射线晶体结构的Ca 2 +ATP：SbtB 7001，显示这种金属离子如何促进额外的稳定与顶点的T-环的相互作用。我们建议，在SbtB 7001中观察到的Ca 2 + ATP诱导的构象变化是重要的SbtB的SbtA活性的变构调节，并与改变腺苷酸水平在光照/黑暗周期是一致的。图形摘要
Cyanobacteria have evolved a suite of enzymes and inorganic carbon (Ci) transporters that improve photosynthetic performance by increasing the localized concentration of CO2 around the primary CO2-fixating enzyme, Rubisco. This CO2-concentrating mechanism (CCM) is highly regulated, responds to illumination/darkness cycles and allows cyanobacteria to thrive under limiting Ci conditions. While the transcriptional control of CCM activity is well understood, less is known about how regulatory proteins might allosterically regulate Ci transporters in response to changing conditions. Cyanobacterial sodium-dependent bicarbonate transporters (SbtAs) are inhibited by PII-like regulatory proteins (SbtBs), with the inhibitory effect being modulated by adenylnucleotides. Here, we used isothermal titration calorimetry to show that SbtB from Cyanobium sp. PCC7001 (SbtB7001) binds AMP, ADP, cAMP and ATP with micromolar-range affinities. X-ray crystal structures of apo- and nucleotide-bound SbtB7001 revealed that while AMP, ADP and cAMP have little effect on the SbtB7001 structure, binding of ATP stabilizes the otherwise flexible T-loop and that the flexible C-terminal C-loop adopts several distinct conformations. We also show that ATP binding affinity is increased ten-fold in the presence of Ca2+ and we present an X-ray crystal structure of Ca2+ATP:SbtB7001 that shows how this metal ion facilitates additional stabilizing interactions with the apex of the T-loop. We propose that the Ca2+ATP-induced conformational change observed in SbtB7001 is important for allosteric regulation of SbtA activity by SbtB and is consistent with changing adenylnucleotide levels in illumination/darkness cycles. GRAPHICAL ABSTRACT
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