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Interactions of L-serine at the active site of serine hydroxymethytransferases: Induction of thermal stability.

Bhaskar, B. and Prakash, V. and Savithri, H. S. and Appaji Rao, N. (1994) Interactions of L-serine at the active site of serine hydroxymethytransferases: Induction of thermal stability. Biochimica et Biophysica Acta, 1209. pp. 40-50.

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Biochimica_et_Biophysica_Acta_(BBA)_-_Protein_Structure_and_Molecular_Enzymology,_Volume_1209,_Issue_1,_16_November_1994,_Pages_40-50.pdf
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Abstract

Serine hydroxymethyltransferase (SHMT), EC 2.1.2.1, exhibits broad substrate and reaction specificity. In addition to cleaving many 3-hydroxyamino acids to glycine and an aldehyde, the enzyme also catalyzed the decarboxylation, transamination and racemization of several substrate analogues of amino acids. To elucidate the mechanism of interaction of substrates, especially L-serine with the enzyme, a comparative study of interaction of L-serine with the enzyme from sheep liver and Escherichia coli, was carried out. The heat stability of both the enzymes was enhanced in the presence of serine, although to different extents. Thermal denaturation monitored by spectral changes indicated an alteration in the apparent T m of sheep liver and E. coli SHMTs from 55 + I°C to 72 + 3°C at 40 mM serine and from 67 + I°C to 72 + I°C at 20 mM serine, respectively. Using stopped flow spectrophotometry k values of (49 + 5). 10 -3 s- 1 and (69 + 7)" 10 -3 S- 1 for sheep liver and E. coli enzymes were determined at 50 mM serine. The binding of serine monitored by intrinsic fluorescence and sedimentation velocity measurements indicated that there was no generalized change in the structure of both proteins. However, visible CD measurements indicated a change in the asymmetric environment of pyridoxal 5'-phosphate at the active site upon binding of serine to both the enzymes. The formation of an external aldimine was accompanied by a change in the secondary structure of the enzymes monitored by far UV-CD spectra. Titration microcalorimetric studies in the presence of serine (8 mM) also demonstrated a single class of binding and the conformational changes accompanying the binding of serine to the enzyme resulted in a more compact structure leading to increased thermal stability of the enzyme.

Item Type: Article
Uncontrolled Keywords: Serine hydroxymethyltransferase; Serine interaction; Thermal stability; Aldimine, internal and external
Subjects: 600 Technology > 08 Food technology > 16 Nutritive value > 02 Amino acids
Divisions: Protein Chemistry and Technology
Depositing User: Food Sci. & Technol. Information Services
Date Deposited: 02 Jun 2011 06:06
Last Modified: 28 Dec 2011 09:41
URI: http://ir.cftri.com/id/eprint/2679

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