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The C-terminal domain of dimeric serine hydroxymethyltransferase plays a key role in stabilization of the quaternary structure and cooperative unfolding of protein: domain swapping studies with enzymes having high sequence identity.
Protein Sci. 2004 Aug; 13(8):2184-95.PS

Abstract

The serine hydroxymethyltransferase from Bacillus subtilis (bsSHMT) and B. stearothermophilus (bstSHMT) are both homodimers and share approximately 77% sequence identity; however, they show very different thermal stabilities and unfolding pathways. For investigating the role of N- and C-terminal domains in stability and unfolding of dimeric SHMTs, we have swapped the structural domains between bs- and bstSHMT and generated the two novel chimeric proteins bsbstc and bstbsc, respectively. The chimeras had secondary structure, tyrosine, and pyridoxal-5'-phosphate microenvironment similar to that of the wild-type proteins. The chimeras showed enzymatic activity slightly higher than that of the wild-type proteins. Interestingly, the guanidium chloride (GdmCl)-induced unfolding showed that unlike the wild-type bsSHMT, which undergoes dissociation of native dimer into monomers at low guanidium chloride (GdmCl) concentration, resulting in a non-cooperative unfolding of enzyme, its chimera bsbstc, having the C-terminal domain of bstSHMT was resistant to low GdmCl concentration and showed a GdmCl-induced cooperative unfolding from native dimer to unfolded monomer. In contrast, the wild-type dimeric bstSHMT was resistant to low GdmCl concentration and showed a GdmCl-induced cooperative unfolding, whereas its chimera bstbsc, having the C- terminal domain of bsSHMT, showed dissociation of native dimer into monomer at low GdmCl concentration and a GdmCl-induced non-cooperative unfolding. These results clearly demonstrate that the C-terminal domain of dimeric SHMT plays a vital role in stabilization of the oligomeric structure of the native enzyme hence modulating its unfolding pathway.

Authors+Show Affiliations

Division of Molecular and Structural Biology, Central Drug Research Institute, Lucknow 226 001, India. bhakuniv@rediffmail.comNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

15273312

Citation

Bhatt, Anant Narayan, et al. "The C-terminal Domain of Dimeric Serine Hydroxymethyltransferase Plays a Key Role in Stabilization of the Quaternary Structure and Cooperative Unfolding of Protein: Domain Swapping Studies With Enzymes Having High Sequence Identity." Protein Science : a Publication of the Protein Society, vol. 13, no. 8, 2004, pp. 2184-95.
Bhatt AN, Khan MY, Bhakuni V. The C-terminal domain of dimeric serine hydroxymethyltransferase plays a key role in stabilization of the quaternary structure and cooperative unfolding of protein: domain swapping studies with enzymes having high sequence identity. Protein Sci. 2004;13(8):2184-95.
Bhatt, A. N., Khan, M. Y., & Bhakuni, V. (2004). The C-terminal domain of dimeric serine hydroxymethyltransferase plays a key role in stabilization of the quaternary structure and cooperative unfolding of protein: domain swapping studies with enzymes having high sequence identity. Protein Science : a Publication of the Protein Society, 13(8), 2184-95.
Bhatt AN, Khan MY, Bhakuni V. The C-terminal Domain of Dimeric Serine Hydroxymethyltransferase Plays a Key Role in Stabilization of the Quaternary Structure and Cooperative Unfolding of Protein: Domain Swapping Studies With Enzymes Having High Sequence Identity. Protein Sci. 2004;13(8):2184-95. PubMed PMID: 15273312.
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TY - JOUR T1 - The C-terminal domain of dimeric serine hydroxymethyltransferase plays a key role in stabilization of the quaternary structure and cooperative unfolding of protein: domain swapping studies with enzymes having high sequence identity. AU - Bhatt,Anant Narayan, AU - Khan,M Yahiya, AU - Bhakuni,Vinod, PY - 2004/7/27/pubmed PY - 2005/2/17/medline PY - 2004/7/27/entrez SP - 2184 EP - 95 JF - Protein science : a publication of the Protein Society JO - Protein Sci VL - 13 IS - 8 N2 - The serine hydroxymethyltransferase from Bacillus subtilis (bsSHMT) and B. stearothermophilus (bstSHMT) are both homodimers and share approximately 77% sequence identity; however, they show very different thermal stabilities and unfolding pathways. For investigating the role of N- and C-terminal domains in stability and unfolding of dimeric SHMTs, we have swapped the structural domains between bs- and bstSHMT and generated the two novel chimeric proteins bsbstc and bstbsc, respectively. The chimeras had secondary structure, tyrosine, and pyridoxal-5'-phosphate microenvironment similar to that of the wild-type proteins. The chimeras showed enzymatic activity slightly higher than that of the wild-type proteins. Interestingly, the guanidium chloride (GdmCl)-induced unfolding showed that unlike the wild-type bsSHMT, which undergoes dissociation of native dimer into monomers at low guanidium chloride (GdmCl) concentration, resulting in a non-cooperative unfolding of enzyme, its chimera bsbstc, having the C-terminal domain of bstSHMT was resistant to low GdmCl concentration and showed a GdmCl-induced cooperative unfolding from native dimer to unfolded monomer. In contrast, the wild-type dimeric bstSHMT was resistant to low GdmCl concentration and showed a GdmCl-induced cooperative unfolding, whereas its chimera bstbsc, having the C- terminal domain of bsSHMT, showed dissociation of native dimer into monomer at low GdmCl concentration and a GdmCl-induced non-cooperative unfolding. These results clearly demonstrate that the C-terminal domain of dimeric SHMT plays a vital role in stabilization of the oligomeric structure of the native enzyme hence modulating its unfolding pathway. SN - 0961-8368 UR - https://www.unboundmedicine.com/medline/citation/15273312/The_C_terminal_domain_of_dimeric_serine_hydroxymethyltransferase_plays_a_key_role_in_stabilization_of_the_quaternary_structure_and_cooperative_unfolding_of_protein:_domain_swapping_studies_with_enzymes_having_high_sequence_identity_ L2 - https://doi.org/10.1110/ps.04769004 DB - PRIME DP - Unbound Medicine ER -