Stability Determination of Intact Humanin-G with Characterizations of Oxidation and Dimerization Patterns.

Autor: Ozgul M; Department of Ophthalmology, Gavin Herbert Eye Institute, University of California Irvine, Irvine, CA 92617, USA., Nesburn AB; Department of Ophthalmology, Gavin Herbert Eye Institute, University of California Irvine, Irvine, CA 92617, USA.; Cedars-Sinai Medical Center, Los Angeles, CA 90048, USA., Nasralla N; IVD Technologies Inc., Santa Ana, CA 92705, USA., Katz B; Department of Chemistry, University of California Irvine, Irvine, CA 92697, USA., Taylan E; Department of Obstetrics and Gynecology, University of California Los Angeles, Los Angeles, CA 90095, USA., Kuppermann BD; Department of Ophthalmology, Gavin Herbert Eye Institute, University of California Irvine, Irvine, CA 92617, USA., Kenney MC; Department of Ophthalmology, Gavin Herbert Eye Institute, University of California Irvine, Irvine, CA 92617, USA.; Department of Pathology and Laboratory Medicine, University of California Irvine, Irvine, CA 92617, USA.
Jazyk: angličtina
Zdroj: Biomolecules [Biomolecules] 2023 Mar 11; Vol. 13 (3). Date of Electronic Publication: 2023 Mar 11.
DOI: 10.3390/biom13030515
Abstrakt: Humanin is the first identified mitochondrial-derived peptide. Humanin-G (HNG) is a variant of Humanin that has significantly higher cytoprotective properties. Here, we describe the stability features of HNG in different conditions and characterize HNG degradation, oxidation, and dimerization patterns over short-term and long-term periods. HNG solutions were prepared in high-performance liquid chromatography (HPLC) water or MO formulation and stored at either 4 °C or 37 °C. Stored HNG samples were analyzed using HPLC and high-resolution mass spectrometry (HRMS). Using HPLC, full-length HNG peptides in HPLC water decreased significantly with time and higher temperature, while HNG in MO formulation remained stable up to 95% at 4 °C on day 28. HNG peptides in HPLC water, phosphate-buffered saline (PBS) and MO formulation were incubated at 37 °C and analyzed at day 1, day 7 and day 14 using HRMS. Concentrations of full-length HNG peptide in HPLC water and PBS declined over time with a corresponding appearance of new peaks that increased over time. These new peaks were identified to be singly oxidized HNG, doubly oxidized HNG, homodimerized HNG, singly oxidized homodimerized HNG, and doubly oxidized homodimerized HNG. Our results may help researchers improve the experimental design to further understand the critical role of HNG in human diseases.
Competing Interests: M.C.K.: Her research on mitochondria has been supported by Discovery Eye Foundation (DEF). She serves as a member of the Board for DEF. The University of California, Irvine has reviewed and approved the terms of this arrangement in accordance with conflicts-of-interest policies. M.C.K.: Collaborations with Allegro, Ophthalmics, LLC and Almon Therapeutics, Inc. B.D.K: CLINICAL RESEARCH: Alcon, Alimera, Allegro, Allergan, Apellis, Clearside, Genentech, GSK, Ionis, jCyte, Novartis, Regeneron, ThromboGenics; CONSULTANT: Alimera, Allegro, Allergan, Cell Care, Dose, Eyedaptic, Galimedix, Genentech, Glaukos, Interface Biologics, jCyte, Novartis, Ophthotech, Regeneron, Revana, Theravance Biopharma.
Databáze: MEDLINE
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