Measurement of Human Surfactant Protein-B Turnover in Vivo from Tracheal Aspirates Using Targeted Proteomics

摘要

We describe a method to measure protein synthesis andncatabolism in humans without prior purification and usenthe method to measure the turnover of surfactant protein-nB (SP-B). SP-B, a lung-specific, hydrophobic proteinnessential for fetal-neonatal respiratory transition, is presentnin only picomolar quantities in tracheal aspirate samplesnand difficult to isolate for dynamic turnover studies usingntraditional in vivo tracer techniques. Using infusion ofn[5,5,5-2H3] leucine and a targeted proteomics method,nwe measured both the quantity and kinetics of SP-Bntryptic peptides in tracheal aspirate samples of symptomaticnnewborn infants. The fractional synthetic raten(FSR) of SP-B measured using the most abundantnproteolytic fragment, a 10 amino acid peptide from thencarboxy-terminus of proSP-B (SPTGEWLPR), from thencirculating leucine pool was 0.035 ( 0.005 h-1, andnthe fractional catabolic rate was 0.044 ( 0.003 h-1.nThis technique permits high-throughput and sensitivenmeasurement of turnover of low abundance proteinsnwith minimal sample preparation.