ePrints@IIScePrints@IISc Home | About | Browse | Latest Additions | Advanced Search | Contact | Help

Structural and functional studies of Bacillus stearothermophilus serine hydroxymethyl transferase: the role of N341, Y60 and F351 in tetrahydrofolate binding.

Pai Verneker, VR and Rajaram , V and Bisht , S and Bhavani , BS and Rao , NA and Murthy , MR and Savithri, HS (2009) Structural and functional studies of Bacillus stearothermophilus serine hydroxymethyl transferase: the role of N341, Y60 and F351 in tetrahydrofolate binding. In: Biochemical Journal, 418 . pp. 635-642.

[img] PDF
4180635.pdf - Published Version
Restricted to Registered users only

Download (852Kb) | Request a copy
Official URL: http://www.biochemj.org/bj/418/bj4180635.htm

Abstract

SHMT (serine hydoxymethyltransferase), a type I pyridoxal 5'-phosphate-dependent enzyme, catalyses the conversion of L-serine and THF (tetrahydrofolate) into glycine and 5,10 -methylene THE SHMT also catalyses several THF-independent side reactions such as cleavage of P-hydroxy amino acids, trans-amination, racemization and decarboxylation. In the present study, the residues Asn(341), Tyr(60) and Phe(351), which are likely to influence THF binding, were mutated to alanine, alanine and glycine respectively, to elucidate the role of these residues in THF-dependent and -independent reactions catalysed by SHMT. The N341A and Y60A bsSHMT (Bacillus stearothermophilus SHMT) mutants were inactive for the THF-dependent activity, while the mutations had no effect on THF-independent activity. However, mutation of Phe(351) to glycine did not have any effect oil either of the activities. The crystal structures of the glycine binary complexes of the mutants showed that N341A bsSHMT forms an external aldimine as in bsSHMT, whereas Y60A and F351G bsSHMTs exist as a Mixture of internal/external aldimine and gem-diamine forms. Crystal structures of all of the three Mutants obtained in the presence of L-allo-threonine were similar to the respective glycine binary complexes. The structure of the ternary complex of F351G bsSHMT with glycine and FTHF (5-formyl THF) showed that the monoglutamate side chain of FTHF is ordered in both the subunits of the asymmetric unit, unlike in the wild-type bsSHMT. The present studies demonstrate that the residues Asn(341) and Tyr(60) are pivotal for the binding of THF/FTHF, whereas Phe(351) is responsible for the asymmetric binding of FTHF in the two subunits of the dimer.

Item Type: Journal Article
Additional Information: Copyright of this article belongs to Portland Press.
Keywords: L-allo-threonine, crystal structure, folate binding, 5-formyl tetrahydrofolate (FTHF), pyridoxal 5′-phosphate, serine hydroxymethyltransferase (SHMT), ternary complex.
Department/Centre: Division of Biological Sciences > Biochemistry
Division of Biological Sciences > Molecular Biophysics Unit
Date Deposited: 14 Dec 2010 06:26
Last Modified: 14 Dec 2010 06:26
URI: http://eprints.iisc.ernet.in/id/eprint/34003

Actions (login required)

View Item View Item