Meeting Abstract
The Pacific whiteleg shrimp, Litopenaeus vannamei, is farmed in environments where O2 fluctuates widely. Its respiratory pigment hemocyanin (Hc) is composed of subunits arranged as n×6-oligomers which have intrinsic O2 binding properties. Shrimp Hc has been reported to have a large and small subunit. However, high throughput RNA-Seq suggests that the large subunit exists as several variants, and that hypoxia might induce changes in their expression. Here we characterize the protein oligomer and subunit compositions of Hc between shrimp exposed to hypoxia for 24h or 14d, compared to normoxic controls. Ratios of 1×6-mer and 2×6-mer Hc were quantified using SEC-HPLC monitored at 280 and 338nm. Oligomers were trypsin-digested to analyze subunit compositions by LC-MS/MS. Proteins were identified by MASCOT searches against combined L. vannamei RNA-Seq assembly, Uniprot penaeid shrimp proteins excluding Hc, and contamination databases. Regardless of hypoxic treatment, L. vannamei Hc existed primarily as 1×6-mer. One Hc small subunit was identified in both 1×6-mer and 2×6-mer, and accounted for 49 and 44% of total Hc normalized spectral counts (NSC) respectively. Three novel large subunit isoforms were identified in both oligomers, and ranged from 14 to 24% of total Hc NSC each. Two novel Hc C-domain variants were identified exclusively in the 2×6-mer at 1% of total Hc NSC. Future use of isobaric tags with LC-MS/MS will more reliably quantify isoform usage in hypoxia relative to normoxia. Understanding how L. vannamei changes Hc in response to hypoxia is important for shrimp aquaculture, and has implications for hypoxic responses in the wild. (NSF IOS-1147008)
