We FRET so You Don’t Have To: New Models of the Lipoprotein Lipase Dimer,Biochemistry

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We FRET so You Don’t Have To: New Models of the Lipoprotein Lipase Dimer
Biochemistry ( IF 2.9 ) Pub Date : 2018-01-05 00:00:00 , DOI: 10.1021/acs.biochem.7b01009
Cassandra K. Hayne ₁ , Hayretin Yumerefendi ₁ , Lin Cao ₁ , Jacob W. Gauer ₂ , Michael J. Lafferty ₁ , Brian Kuhlman _{1,

3} , Dorothy A. Erie _{2,

3} , Saskia B. Neher ₁

Affiliation

Lipoprotein lipase (LPL) is a dimeric enzyme that is responsible for clearing triglyceride-rich lipoproteins from the blood. Although LPL plays a key role in cardiovascular health, an experimentally derived three-dimensional structure has not been determined. Such a structure would aid in understanding mutations in LPL that cause familial LPL deficiency in patients and help in the development of therapeutic strategies to target LPL. A major obstacle to structural studies of LPL is that LPL is an unstable protein that is difficult to produce in the quantities needed for nuclear magnetic resonance or crystallography. We present updated LPL structural models generated by combining disulfide mapping, computational modeling, and data derived from single-molecule Förster resonance energy transfer (smFRET). We pioneer the technique of smFRET for use with LPL by developing conditions for imaging active LPL and identifying positions in LPL for the attachment of fluorophores. Using this approach, we measure LPL–LPL intermolecular interactions to generate experimental constraints that inform new computational models of the LPL dimer structure. These models suggest that LPL may dimerize using an interface that is different from the dimerization interface suggested by crystal packing contacts seen in structures of pancreatic lipase.

中文翻译：

我们进行FRET，因此您不必：脂蛋白脂肪酶二聚体的新模型

脂蛋白脂肪酶（LPL）是一种二聚酶，负责从血液中清除富含甘油三酸酯的脂蛋白。尽管LPL在心血管健康中起着关键作用，但尚未确定实验得出的三维结构。这样的结构将有助于理解引起患者家族性LPL缺乏的LPL突变，并有助于开发靶向LPL的治疗策略。LPL结构研究的主要障碍是LPL是一种不稳定的蛋白质，难以产生核磁共振或晶体学所需的数量。我们介绍了通过结合二硫键映射，计算模型和从单分子Förster共振能量转移（smFRET）得出的数据生成的更新的LPL结构模型。我们通过开发对有源LPL成像的条件并确定LPL中荧光团的附着条件，开创了可与LPL一起使用的smFRET技术的先河。使用这种方法，我们测量LPL-LPL的分子间相互作用，以产生实验性约束条件，从而为LPL二聚体结构的新计算模型提供依据。这些模型表明，LPL可能使用与胰脂肪酶结构中所见的晶体堆积接触所暗示的二聚化界面不同的二聚化界面。

更新日期：2018-01-05

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