Functional mode of NtHSP17.6, a cytosolic small heat-shock protein fromNicotiana tabacum

Abstract

Small heat-shock proteins (sHsps) are ubiquitous stress proteins with molecular chaperone activity. They share characteristic homology with the @-crystallin protein of the mammalian eye lens as well as being ATP-independent in their chaperone activity. We isolated a clone for a cytosolic class ! sHsp, NtHSP17.6, from Nicotiana tabacum, and analyzed its functional mode for such activity. Following its transformation into Escherichia coli and its over-expression, NtHSP17.6 was purified and examined in vitro. This purified NtHSP17.6 exhibited typical chaperone activity in a lightscattering test. It was enable to protect a model substrate, firefly luciferase, from heat-induced aggregation. Nondenaturing PAGE showed that NtHSP17.6 formed a dodecamer in its native conformation, and was bound to its substrate under heat stress. A labeling test with bis-ANS indicated that this binding might be linked to newly exposed hydrophobic sites of the NtHSP17.6 complexes during heat shock. Based on these data, we suggest that NtHSP17.6 is a molecular chaperone that functions as a dodecamer in a heat-induced manner.

Model Chaperones: Small Heat Shock Proteins from Plants

“…Both Ta16.9 and Ps18.1 are dodecamers in solution at room temperature, consistent with the Ta16.9 crystal structure, and all available data indicate that sHSPs in the plant class I family most likely all assemble into dodecamers Kirschner et al 2000 ;Tiroli and Ramos 2007 ;Yoon et al 2005 ). It has been suggested that sHSP oligomers act as reservoirs of the active dimeric units of the chaperone van Montfort et al 2001b ).…”

Molecular characterization of theCapsicum annuum RING zinc finger protein 1 (CaRZFP1) gene induced by abiotic stresses

“…Although high temperature is a major abiotic stress that severely damages crop productivity, most previous research on plant responses at the molecular level has centered on heat shock proteins (HSPs) (Cho and Hong, 2004;Yoon et al, 2005) and heat shock transcription factors (HSFs). HSFs are constitutively present and, under non-stressed conditions, they exist as monomer forms that usually bind to heat shock protein 70 (HSP70).…”

A novel J8 domain gene,IbJ8, inIpomoea batatas (L). Lam.

“…In addition, a large number of proteins containing a single J-domain are combined with other domains distinct from those in E. coil DnaJ (Kelley, 1998). The J-domain, which is the definitive region for all DnaJ proteins, interacts with Hsp70 proteins and stimulates the Hsp70 ATPase activity necessary for stable binding of Hsp70 to its protein substrate (Bukau and Horwich, 1998;Cho and Hong, 2004;Yoon et a[., 2005). Membrane-bound forms of DnaJ proteins invariably contain this functional domain as well as numerous transmembrane regions (Kelley, 1998).…”