Dataset Information

Elucidation of Lipid Binding Sites on Lung Surfactant Protein A Using X-ray Crystallography, Mutagenesis, and Molecular Dynamics Simulations.

ABSTRACT: Surfactant protein A (SP-A) is a collagenous C-type lectin (collectin) that is critical for pulmonary defense against inhaled microorganisms. Bifunctional avidity of SP-A for pathogen-associated molecular patterns (PAMPs) such as lipid A and for dipalmitoylphosphatidylcholine (DPPC), the major component of surfactant membranes lining the air-liquid interface of the lung, ensures that the protein is poised for first-line interactions with inhaled pathogens. To improve our understanding of the motifs that are required for interactions with microbes and surfactant structures, we explored the role of the tyrosine-rich binding surface on the carbohydrate recognition domain of SP-A in the interaction with DPPC and lipid A using crystallography, site-directed mutagenesis, and molecular dynamics simulations. Critical binding features for DPPC binding include a three-walled tyrosine cage that binds the choline headgroup through cation-? interactions and a positively charged cluster that binds the phosphoryl group. This basic cluster is also critical for binding of lipid A, a bacterial PAMP and target for SP-A. Molecular dynamics simulations further predict that SP-A binds lipid A more tightly than DPPC. These results suggest that the differential binding properties of SP-A favor transfer of the protein from surfactant DPPC to pathogen membranes containing appropriate lipid PAMPs to effect key host defense functions.

SUBMITTER: Goh BC

PROVIDER: S-EPMC5663190 | biostudies-literature | 2016 Jul

REPOSITORIES: biostudies-literature

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Publications

Elucidation of Lipid Binding Sites on Lung Surfactant Protein A Using X-ray Crystallography, Mutagenesis, and Molecular Dynamics Simulations.

Goh Boon Chong BC Wu Huixing H Rynkiewicz Michael J MJ Schulten Klaus K Seaton Barbara A BA McCormack Francis X FX

Biochemistry 20160621 26

Surfactant protein A (SP-A) is a collagenous C-type lectin (collectin) that is critical for pulmonary defense against inhaled microorganisms. Bifunctional avidity of SP-A for pathogen-associated molecular patterns (PAMPs) such as lipid A and for dipalmitoylphosphatidylcholine (DPPC), the major component of surfactant membranes lining the air-liquid interface of the lung, ensures that the protein is poised for first-line interactions with inhaled pathogens. To improve our understanding of the mot ...[more]

PMID: 27324153

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Project description:Alveolar macrophages (AMs) are CD44 expressing cells that reside in the alveolar space where they maintain lung homeostasis by serving critical roles in immunosurveillance and lipid surfactant catabolism. AMs lacking CD44 are unable to bind the glycosaminoglycan, hyaluronan, which compromises their survival and leads to reduced numbers of AMs in the lung. Using RNA sequencing, lipidomics and multiparameter flow cytometry, we demonstrate that CD44-/- mice have impaired AM lipid homeostasis and increased surfactant lipids in the lung. CD44-/- AMs had increased expression of CD36, a lipid scavenger receptor, as well as increased intracellular lipid droplets, giving them a foamy appearance. RNA sequencing revealed the differential expression of genes associated with lipid efflux and metabolism in CD44-/- AMs. Lipidomic analysis showed increased lipids in both the supernatant and cell pellet extracted from the bronchoalveolar lavage of CD44-/- mice. Phosphatidylcholine species, cholesterol, oxidized phospholipids and levels of reactive oxygen species (ROS) were increased in CD44-/- AMs. Oxidized phospholipids were more cytotoxic to CD44-/- AMs and induced greater lung inflammation in CD44-/- mice. Reconstitution of CD44+/+ mice with CD44-/- bone marrow as well as adoptive transfer of CD44-/- AMs into CD44+/+ mice showed that lipid accumulation in CD44-/- AMs occurred irrespective of the lung environment, suggesting a cell intrinsic defect. Administration of colony stimulating factor 2 (CSF-2), a critical factor in AM development and maintenance, increased AM numbers in CD44-/- mice and decreased phosphatidylcholine levels in the bronchoalveolar lavage, but was unable to decrease intracellular lipid accumulation in CD44-/- AMs. Peroxisome proliferator-activated receptor gamma (PPARγ), downstream of CSF-2 signaling and a regulator of lipid metabolism, was reduced in the nucleus of CD44-/- AMs, and PPARγ inhibition in normal AMs increased their lipid droplets. Thus, CD44 deficiency causes defects in AMs that lead to abnormal lipid accumulation and oxidation, which exacerbates oxidized lipid-induced lung inflammation. Collectively, these findings implicate CD44 as a regulator of lung homeostasis and inflammation.

Dataset Information

Elucidation of Lipid Binding Sites on Lung Surfactant Protein A Using X-ray Crystallography, Mutagenesis, and Molecular Dynamics Simulations.

Publications

Elucidation of Lipid Binding Sites on Lung Surfactant Protein A Using X-ray Crystallography, Mutagenesis, and Molecular Dynamics Simulations.

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