Table 2.
Summary of peptide production and purification schemes
| Target peptide | Final yield of respective schemes (μg/mg wet cell pellet)a | ||
|---|---|---|---|
| Scheme 1A | Scheme 1B | Scheme 2 | |
| Step 1: Expression of fusion protein Peptide-Intein-ELK16 | Step 1: Expression of fusion protein Peptide-Intein-ELK16 | Step 1: Expression of fusion protein Trx-Peptide-Intein-ELK16 | |
| Step 2: Intein-mediated cleavage | Step 2: Intein-mediated cleavage | Step 2: Intein-mediated cleavage | |
| Step 3: RP-HPLC | Step 3: Refolding | Step 3: Enterokinase cleavage | |
| Step 4: RP-HPLC | Step 4: RP-HPLC | ||
| GLP-1 | 0.8 (20.8 %)b | – | 1.1 (25.2 %) |
| BNP | No expression | – | 1.8 (42.2 %) |
| Ex-4 | No expression | – | 0.3 (26.5 %) |
| IGF-1α | Insoluble | 0.3 (3.6 %) | Unwanted enterokinase site |
| CCL5 | Insoluble | 0.1 (1.2 %) | Insoluble |
| SDF-1α | Insoluble | 0.4 (5.8 %) | 0.2 (2.0 %) |
| Leptin | Insoluble | 0.6 (3.4 %) | Insoluble |
aYield of target peptide after fine purification by RP-HPLC from LB culture with wet cell weight of 2.66 ± 0.99 mg/mL, quantified by the Pierce® bicinchoninic acid (BCA) assay kit
bThe final recovery after RP-HPLC purification listed in parentheses and calculated by dividing the product peptide yield by the theoretical value from aggregate yield in Table 1