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Biochemistry. 1986 Oct 7;25(20):5998-6004.

Preparation and properties of porcine relaxin derivatives shortened at the amino terminus of the A chain.

Abstract

Porcine relaxins shortened at the N terminus of the A chain were produced after protection of all amino groups with the base-labile [[(methylsulfonyl)ethyl]oxy]carbonyl (Msc) protecting group. The first two amino acids were removed by cyanogen bromide digestion whereby simultaneously a free alpha-amino group was generated in position A3. The resulting des-ArgA1,MetA2-N epsilon A7,N epsilon A16,N epsilon B8-tris [[[(methylsulfonyl)ethyl]oxy]carbonyl]relaxin. was further shortened by preparative Edman degradation. The shortest derivative obtained was des-ArgA1,MetA2,ThrA3,LeuA4,SerA5,GluA6 -N epsilon A7,N epsilon A16,N epsilon B8-tris[[[(methylsulfonyl)ethyl]oxy]carbonyl]relaxin. The deprotection of the derivatives in alkaline media resulted in crude des-A(1-2)- to des-A(1-6)-relaxins, which were subsequently purified by gel filtration on Sephadex G-50 superfine followed by either ion exchange chromatography on CM-cellulose at pH 5.1 or high-performance liquid chromatography on reversed-phase columns. During the CNBr digest, a side product was isolated that was identified as the corresponding homoserine ( [HseA2]relaxin) derivative. Shortened relaxin derivatives and [HseA2]relaxin were characterized by reversed-phase chromatography, electrophoresis, end-group determination, and amino acid composition. Circular dichroism studies revealed a distinct change in the structure of relaxins that were shortened by three and more amino acid residues. In the mouse interpubic ligament assay, des-A(1-2)-relaxin and [HseA2]relaxin were fully biologically active while the bioactivity of des-A(1-3)-relaxin dropped to about 50%. Relaxins shortened by four and more amino acid residues were biologically inactive.(ABSTRACT TRUNCATED AT 250 WORDS).

PMID:
3790502
DOI:
10.1021/bi00368a025
[Indexed for MEDLINE]

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