Open AccessThe proline‐activating activity of the multienzyme gramicidin S synthetase 2 can be recovered on a 115‐kDa tryptic fragment
Author(s) -
SKARPEID HansJacob,
ZIMMER TrineLise,
SHEN Beifen,
DÖHREN Hans
Publication year - 1990
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1990.tb15346.x
Subject(s) - gramicidin s , proteolysis , trypsin , gramicidin , biochemistry , molecular mass , cleavage (geology) , chemistry , recombinant dna , enzyme , mutant , proline , biology , stereochemistry , amino acid , gene , paleontology , membrane , fracture (geology)
The multienzyme gramicidin S synthetase 2 was treated with trypsin to obtain fragments capable of activating proline. Three different active fragments were detected. The course of proteolysis was simulated by using a concentration range of trypsin; the cleavage pattern indicated that one of the fragments was particularly stable. This fragment was purified and shown to have a molecular mass of 115 kDa. It was compared chromatographically, by SDS/PAGE, and enzymatically to a Pro‐activating fragment produced by a gramicidin‐S‐negative mutant. It can be concluded that the proteolytic fragment represents a structure which is contained on a continuous part of the polypeptide chain of gramicidin S synthetase 2 and has a relatively compact structure. This provides evidence that the multienzyme gramicidin S synthetase 2 is, at least in part, constructed from functional domains. An approach towards extending these studies to other parts of the gramicidin S synthetase 2 molecule has also been devised. This work complements recombinant DNA studies in the area, providing stable functional fragments.