Purification of native surfactant protein SP-A from pooled amniotic fluid and bronchoalveolar lavage.
Surfactant protein SP-A is a hydrophilic glycoprotein, similar to SP-D, which plays an important role in pulmonary surfactant homeostasis and innate immunity. SP-A is actively expressed in the alveolar type II cells and Clara cells. Their basic structure consists of triple-helical collagen region and a C-terminal carbohydrate recognition domain (CRD). By binding to the infectious microbes, SP-A (like SP-D) are involved in pathogen opsonization and agglutination and subsequent clearance of the microorganism, via recruitment of phagocytic cells via receptors for the collagen region. SP-A has also been localized at extra-pulmonary sites such as salivary epithelium, amniotic fluid, prostate glands, and semen. The presence of SP-A in fetal and maternal tissue and amniotic fluid suggests it is involved in pregnancy and labor. Native SP-A can be purified from amniotic fluid and bronchiolar lavage fluid (BALF) via affinity chromatography. In addition, we also report here a procedure to express and purify a recombinant form of trimeric CRD in Escherichia coli. The availability of highly pure native SP-A and CRD region can be central to studies that examine the diverse roles that SP-A play in surfactant homeostasis, pulmonary infection and inflammation and pregnancy.
Centre for Infection, Immunity and Disease Mechanisms, Biosciences, School of Health Sciences and Social Care, Brunel University, London, UK.
Najmunisa
N
Karbani
Eswari
E
Dodagatta-Marri
Asif S
AS
Qaseem
Priyaa
P
Madhukaran
Patrick
P
Waters
Anthony G
AG
Tsolaki
Taruna
T
Madan
Uday
U
Kishore
D000653:Amniotic Fluid / Q000737:chemistry*; D061249:Batch Cell Culture Techniques; D001992:Bronchoalveolar Lavage Fluid / Q000737:chemistry*; D002846:Chromatography, Affinity; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism; D015870:Gene Expression; D006801:Humans; D011489:Protein Denaturation; D058849:Protein Refolding; D037662:Pulmonary Surfactant-Associated Protein A / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification
D037662:Pulmonary Surfactant-Associated Protein A; D011994:Recombinant Proteins