Home|Journals|Articles by Year|Audio Abstracts
 

Original Article

J App Pharm Sci. 2021; 11(12): 183-195


Expression of human serum albumin in Pichia pastoris protease-deficient host and conjugation with gadolinium-diethylenetriamine pentaacetate for application as a contrast agent

Shabarni Gaffar, Rina Anggraeni, Mia Tria Novianti, Safri Ishmayana, Ukun M. S. Soedjanaatmadja.




Abstract

Human serum albumin (HSA) is a plasma protein with a molecular weight of 66.5 kDa, which is most widely used in the pharmaceutical and clinical fields. One application of HSA in a drug delivery system is to increase the relaxation time on magnetic resonance imaging. The combination of contrast agent, gadolinium-diethylenetriamine pentaacetate (Gd-DTPA), with HSA can produce better imaging. This study aims to express the recombinant HSA (rHSA) in the Pichia pastoris protease-deficient host, SMD1168, purification using chromatographic techniques and conjugation with Gd-DTPA. Optimization of rHSA expression by P. pastoris was done by varying the methanol inducer, 0.75%, 1.0%, 1.25%, and 1.5%, as well as characterization by sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE). Furthermore, the rHSA was purified using gel filtration, characterization by Size Exclusion Chromatography- High Performance Liquid Chromatography (SEC-HPLC), and conjugation with Gd-DTPA. The rHSA (~66.5 kDa) was characterized from broth medium of P. pastoris SMD1168. The optimum conditions for rHSA expression were 1.5% methanol at 48 hours incubation, which resulted in 0.44 g/l of rHSA. The rHSA with 81.84% purity was obtained by gel filtration, and conjugation with Gd-DTPA produced high-purity Gd-DTPA-rHSA, demonstrated by a single peak (rt = 10.493 minutes). The protease-deficient P. pastoris was successfully used to express intact rHSA and the rHSA was successfully conjugated to Gd-DTPA with high purity.

Key words: recombinant Human Serum Albumin, P. pastoris protease-deficient host, Gadolinium-Diethylene Triamine Penta Acetate-HSA.






Full-text options


Share this Article


Online Article Submission
• ejmanager.com




ejPort - eJManager.com
Refer & Earn
JournalList
About BiblioMed
License Information
Terms & Conditions
Privacy Policy
Contact Us

The articles in Bibliomed are open access articles licensed under Creative Commons Attribution 4.0 International License (CC BY), which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons licence, and indicate if changes were made. To view a copy of this licence, visit http://creativecommons.org/licenses/by/4.0/.