Recombinant Human Relaxin Characterization

Introduction to recombinant human Relaxin

Relaxin, a protein hormone predominantly recognized for its significant function in the reproductive biology across various species. This unique protein orchestrates fluctuations in organ structure amid pregnancy and parturition by manipulating the reconfiguration of connective tissues within targeted organs. Moreover, relaxin is known to govern several biological responses in reproductive tissues in gestating animals, a topic we shall delve into, more extensively, in later sections.

Fig 1. Key mechanisms of the cardiovascular complications of diabetes and proposed protective actions of relaxin to target these pathways (indicated by red cross). (Ng, H. H., et al. 2018)

One of the potentially critical applications of relaxin as a therapeutic entity involves suppression of premature labor whilst inducing the process of cervical ripening prior to parturition. Numerous primitive and somewhat unregulated human clinical studies have been conducted, involving the use of porcine relaxin extracted from structures such as the corpus lutea and ovaries.

Primary Structure—Peptide Map

Recombinant human relaxin and its native relaxin obtained from ectopic pregnancy corpus luteum were chemically characterized using tryptic peptide mapping and mass spectrometry. Due to the limited number of native samples, it was not possible to perform a complete chemical analysis or mass collection of the entire relaxin molecule. This problem was addressed by calculating the masses of the individual A and B chains after reduction. The mass of the isolated native A chain matches the predicted mass of the cDNA-derived primary structure, taking into account the pyroglutamate amino terminus of the A chain.

Fig 2. Structure of native and manufactured human relaxin. (Teichman, S. L., et al. 2008)

It is challenging to determine the actual size of the B chain because of the unpredictable protein hydrolysis processing sites in the procreatin sequence. By analogy with the processing sites of porcine and rat prorexins, the mass of the 33-amino acid B chain was predicted to be approximately 3798.6. However, the mass of the independent natural 29-amino acid B chain was detected to be significantly smaller at 3319.9.

Oxidation in the Presence of Light

Human relaxin is a protein with a B-chain length of 33 amino acids, which is degraded by strong light irradiation. This was achieved by applying reversed-phase high-performance liquid chromatography (RP-HPLC) and mass spectrometry.

Fig 3. Reversed-phase HPLC analysis of synthetic human relaxin exposed to 3600 foot-candles of light for 0, 10, and 14 days. (Pearlman, Rodney, and Y. John Wang, eds. 1996)

Upon exposure to light intensity equating to 3600 foot-candles for a duration of 17 days, the RP-HPLC analysis indicated notable alterations in the structure of relaxin. Further examination entailing a tryptic peptide analysis, executed both pre and post intense light exposure, documented alterations in the T6 and T9 tryptic peptides of the relaxin molecule.

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References

1. Ng, H. H., et al. Relaxin as a Therapeutic Target for the Cardiovascular Complications of Diabetes. Front Pharmacol. 2018; 9: 501.
2. Teichman, S. L., et al. Relaxin, a pleiotropic vasodilator for the treatment of heart failure. Heart Failure Reviews. 2008, 14(4), 321–329.
3. Pearlman, Rodney, and Y. John Wang, eds. Formulation, characterization, and stability of protein drugs. Vol. 9. Springer Science & Business Media. 1996.

For research use only, not intended for any clinical use.