Substrate specificities of cyclosporin synthetase and peptolide SDZ 214-103 synthetase. Comparison of the substrate specificities of the related multifunctional polypeptides.
The recently discovered multifunctional polypeptide cyclosporin synthetase is capable of synthesizing the cyclic undecapeptide cyclosporin A in a batch reaction. Substrates are the unmethylated constitutive amino acids of cyclosporin A. Exchange of one or more of these by various amino acids gives a picture of the substrate specificity of the enzyme in vitro, which is different from the known picture obtained by in vivo studies. The uncommon amino acid butenylmethylthreonine in position 1 of the cyclosporin ring can be exchanged by an unexpected large spectrum of different amino acids, showing a great flexibility of this site. Position 2, on the other hand, which shows the greatest variability in vivo, has an only slightly lower specificity in vitro. Position 3 has a very high degree of specificity; positions 4, 6, 7, 9, and 10 have marginally less. The variability of positions 5 and 11 is moderate, whereas position 8 shows only low substrate specificity in vitro. In general, most sites of SDZ 214-103 synthetase appear to be more specific than those of cyclosporin synthetase. Site 11 has nearly identical substrate specificity compared with that of cyclosporin synthetase. The D-2-hydroxy acid position (position 8) can be occupied by a large spectrum of substrates varying from D-lactic acid to D-2-hydroxyisocaproic acid. Within the limits of the present data, the addition of further functional groups to the D-2-hydroxy acid moiety are apparently not tolerated by the enzyme.
Lawen
A
A
Institut für Biochemie und Molekulare Biologie, Technische Universität Berlin, Federal Republic of Germany.
Traber
R
R
D000595:Amino Acid Sequence; D000596:Amino Acids / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D002855:Chromatography, Thin Layer; D016572:Cyclosporine / Q000096:biosynthesis; D003524:Cyclosporins / Q000096:biosynthesis* / Q000737:chemistry / Q000302:isolation & purification; D003904:Mitosporic Fungi / Q000201:enzymology*; D008969:Molecular Sequence Data; D015394:Molecular Structure; D009097:Multienzyme Complexes / Q000378:metabolism*; D010453:Peptide Synthases / Q000378:metabolism*; D013379:Substrate Specificity
D000596:Amino Acids; D003524:Cyclosporins; D009097:Multienzyme Complexes; C048525:cyclosporin A synthetase; C070510:SDZ 214-103; C083273:peptolide SDZ 214-103 synthetase; D016572:Cyclosporine; D010453:Peptide Synthases