Leveraging host-imposed metal starvation to elucidate the molecular and environmental factors that dictate metal utilization by the iron/manganese superoxide dismutase superfamily

利用宿主施加的金属饥饿来阐明决定铁/锰超氧化物歧化酶超家族利用金属的分子和环境因素

基本信息

批准号：
10617269
负责人：
Thomas Everett Kehl-Fie
金额：
$ 57.31万
依托单位：
UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-05-19 至 2026-04-30
项目状态：
未结题

项目摘要

Project Summary/Abstract: Superoxide is a toxic molecule that all organisms exposed to oxygen must cope with. This is particularly true for pathogenic microbes, as the host harnesses the toxic properties of superoxide to combat invaders via the oxidative burst. To detoxify superoxide, nearly all forms of life, including strict anaerobes, produce superoxide dismutase (SOD). Convergent evolution has led to the development of three independent SOD families, all of which are dependent on metals for function. The most widely distributed family of SODs are those which depend on iron (Fe) or manganese (Mn) for function. Members of the Fe/Mn superfamily are present in eukaryotes, archaea, and bacteria. Despite over forty years of study, it is not possible to predict accurately the metal utilized by members of the Fe/Mn superfamily of SODs. Difficulties in predicting metalloprotein metal utilization are not confined to the Fe/Mn SOD superfamily but also occur with other classes of metalloenzymes. This deficiency is driven by relatively low levels of protein sequence identity amongst SODs from different organisms that utilize the same metal cofactor. Additionally, the environmental and molecular factors that dictate the metal used by members of this protein superfamily are also unknown. Members of the Fe/Mn SOD superfamily are canonically thought to use either Fe or Mn, but not both, as a cofactor. This idea arose despite early investigations that identified Fe/Mn SOD family members that are active with both Fe and Mn. The ability of these “cambialistic” SODs (able to use either Fe or Mn as a catalytic cofactor) was dismissed as a quirk of chemistry. At the time, it was thought that intracellular metal concentrations did not change enough to alter the metal bound by a SOD. S. aureus possesses two superoxide dismutases, SodA and SodM, which are ~75% identical. Initially, both SODs were reported to be Mn-dependent. During infection, the host restricts the availability of Mn and inactivates Mn-dependent SODs via the Mn-binding immune protein calprotectin. Recent work discovered that SodM critically contributes to the ability of S. aureus to maintain a defense against oxidative stress when Mn-starved, both in culture and during infection, while SodA is important when Mn is freely available. Biochemical analyses revealed that SodM is not strictly Mn-dependent but is instead cambialistic, and the ability to use Fe enables it to promote resistance to oxidative stress when S. aureus is Mn-limited by the host. These observations support a physiological role for cambialism and the hypothesis that metal availability shapes the repertoire of SODs possessed by an organism. The experiments in this proposal will evaluate this hypothesis and elucidate the molecular features that dictate metal utilization in the Fe/Mn SOD superfamily. Aim I: Elucidate the molecular features that dictate metal utilization of Fe/Mn SOD superfamily members. Aim II: Determine if environmental metal availability promotes retention of a metal-specific and cambialistic SOD by S. aureus. Aim III: Elucidate the broader contribution of cambialistic SODs to maintaining a defense against superoxide.

项目摘要/摘要：超氧化物是一个有毒分子，所有暴露于氧气的生物必须应对。尤其如此病原微生物，因为宿主利用超氧化物的有毒特性通过氧化爆发。为了排毒超氧化物，几乎所有形式的生命，包括严格的厌氧菌，都会产生超氧化物歧义酶（SOD）。融合进化导致了三个独立的草皮家庭的发展取决于金属的功能。分布最广泛的草皮家族是在铁（Fe）或锰（MN）上进行功能。 FE/MN超家族的成员存在于真核生物中，古细菌和细菌。尽管研究了40多年，但仍无法准确预测使用的金属由SOD的Fe/Mn超家族成员。预测金属蛋白金属利用率的困难不是局限于Fe/Mn Sod超家族，但也与其他类别的金属酶一起发生。这种缺陷是来自利用不同生物的草皮中的蛋白质序列身份相对较低的驱动相同的金属辅因子。此外，决定了由金属使用的环境和分子因素该蛋白质超家族的成员也是未知的。 Fe/Mn Sod超家族的成员是典型的被认为使用Fe或Mn，但不是两者都作为辅助因子。这个想法引起了拼命的早期调查鉴定出具有FE和MN活跃的FE/MN SOD家族成员。这些“坎比主义”的能力 SOD（能够使用Fe或Mn作为催化辅助因子）被视为化学怪癖。当时人们认为细胞内金属浓度不足以改变由草皮结合的金属。金黄色葡萄球菌具有两个超氧化物歧化酶，苏打和SODM，约75％相同。最初，两个据报道，草种依赖于Mn。在感染期间，宿主限制了MN的可用性并灭活 Mn结合免疫蛋白钙染色素通过Mn依赖性的SOD。最近的工作发现SODM 批判性地有助于金黄色葡萄球菌在MN饥饿时保持防御氧化应激的能力，在培养和感染期间，当MN免费提供时，苏打水很重要。生化分析揭示了SODM并非严格取决于Mn的依赖性，而是坎比式的，并且使用FE的能力可以使它当金黄色葡萄球菌被宿主限制时，促进氧化应激的抗性。这些观察支持金属可用性塑造了SOD曲目由生物体拥有。该提案中的实验将评估这一假设并阐明分子特征决定了Fe/Mn Sod超家族中金属利用率。目的I：阐明决定了Fe/Mn Sod超家族中金属利用率的分子特征。决定金属利用Fe/Mn Sod超家族成员的特征。 AIM II：确定环境是否金属的可用性促进金黄色葡萄球菌对金属特异性和坎比式SOD的保留。 AIM III：阐明卫生草皮的更广泛的贡献是维持对超氧化物的防御。