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Journal of Virology, January 2006, p. 412-425, Vol. 80, No. 1
0022-538X/06/$08.00+0     doi:10.1128/JVI.80.1.412-425.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.

N- and C-Terminal Cooperation in Rotavirus Enterotoxin: Novel Mechanism of Modulation of the Properties of a Multifunctional Protein by a Structurally and Functionally Overlapping Conformational Domain

M. R. Jagannath,1 M. M. Kesavulu,2 R. Deepa,1 P. Narayan Sastri,1 S. Senthil Kumar,1 K. Suguna,2 and C. Durga Rao1*

Department of Microbiology and Cell Biology,1 Molecular Biophysics Unit, Indian Institute of Science, Bangalore 560012, India2

Received 20 May 2005/ Accepted 4 October 2005

Rotavirus NSP4 is a multifunctional endoplasmic reticulum (ER)-resident nonstructural protein with the N terminus anchored in the ER and about 131 amino acids (aa) of the C-terminal tail (CT) oriented in the cytoplasm. Previous studies showed a peptide spanning aa 114 to 135 to induce diarrhea in newborn mouse pups with the 50% diarrheal dose approximately 100-fold higher than that for the full-length protein, suggesting a role for other regions in the protein in potentiating its diarrhea-inducing ability. In this report, employing a large number of methods and deletion and amino acid substitution mutants, we provide evidence for the cooperation between the extreme C terminus and a putative amphipathic {alpha}-helix located between aa 73 and 85 (AAH73-85) at the N terminus of {Delta}N72, a mutant that lacked the N-terminal 72 aa of nonstructural protein 4 (NSP4) from Hg18 and SA11. Cooperation between the two termini appears to generate a unique conformational state, specifically recognized by thioflavin T, that promoted efficient multimerization of the oligomer into high-molecular-mass soluble complexes and dramatically enhanced resistance against trypsin digestion, enterotoxin activity of the diarrhea-inducing region (DIR), and double-layered particle-binding activity of the protein. Mutations in either the C terminus, AAH73-85, or the DIR resulted in severely compromised biological functions, suggesting that the properties of NSP4 are subject to modulation by a single and/or overlapping highly sensitive conformational domain that appears to encompass the entire CT. Our results provide for the first time, in the absence of a three-dimensional structure, a unique conformation-dependent mechanism for understanding the NSP4-mediated pleiotropic properties including virus virulence and morphogenesis.

* Corresponding author. Mailing address: Department of Microbiology and Cell Biology, Indian Institute of Science, Bangalore 560012, India. Phone: 91-80-23602149. Fax: 91-80-23602697. E-mail: cdr{at}mcbl.iisc.ernet.in.

Journal of Virology, January 2006, p. 412-425, Vol. 80, No. 1
0022-538X/06/$08.00+0     doi:10.1128/JVI.80.1.412-425.2006
Copyright © 2006, American Society for Microbiology. All Rights Reserved.



This article has been cited by other articles:

  • Seo, N.-S., Zeng, C. Q.-Y., Hyser, J. M., Utama, B., Crawford, S. E., Kim, K. J., Hook, M., Estes, M. K. (2008). Inaugural Article: Integrins {alpha}1{beta}1 and {alpha}2{beta}1 are receptors for the rotavirus enterotoxin. Proc. Natl. Acad. Sci. USA 105: 8811-8818 [Abstract] [Full Text]  
  • Rajasekaran, D., Sastri, N. P., Marathahalli, J. R., Indi, S. S., Pamidimukkala, K., Suguna, K., Rao, C. D. (2008). The flexible C terminus of the rotavirus non-structural protein NSP4 is an important determinant of its biological properties. J. Gen. Virol. 89: 1485-1496 [Abstract] [Full Text]  


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