Subunit exchange, conformational stability, and chaperone-like function of the small heat shock protein 16.5 from Methanococcus jannaschii.
Hsp16.5, isolated from the hyperthermophilic Archaea Methanococcus jannaschii, is a member of the small heat-shock protein family. Small Hsps have 12- to 42-kDa subunit sizes and have sequences that are conserved among all organisms. The recently determined crystal structure of Hsp16.5 indicates that it consists discretely of 24 identical subunits. Using fluorescence resonance energy transfer, we show that at temperatures above 60 degrees C, the subunits of MjHsp16.5 freely and reversibly exchange with a rate constant of exchange at 68 degrees C of 0.067 min(-1). The subunit exchange reactions were strongly temperature-dependent, similar to the exchange reactions of the alpha-crystallins. The exchange reaction was specific to MjHsp16.5 subunits, as other sHsps such as alpha-crystallin were not structurally compatible and could not integrate into the MjHsp16.5 oligomer. In addition, we demonstrate that at temperatures as high as 70 degrees C, MjHsp16.5 retains its multimeric structure and subunit organization. Using insulin and alpha-lactalbumin as model target proteins, we also show that MjHsp16.5 at 37 degrees C is a markedly inefficient chaperone compared with other sHsps with these substrates. The results of this study support the hypothesis that MjHsp16.5 has a dynamic quaternary structure at temperatures that are physiologically relevant to M. jannaschii.
Bova
Michael P
MP
Jules Stein Eye Institute, UCLA School of Medicine, Los Angeles, California 90095, USA.
Huang
Qingling
Q
Ding
Linlin
L
Horwitz
Joseph
J
D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000378:metabolism; D000818:Animals; D019843:Archaeal Proteins / Q000737:chemistry / Q000378:metabolism*; D002942:Circular Dichroism; D004229:Dithiothreitol / Q000737:chemistry; D031541:Fluorescence Resonance Energy Transfer; D006360:Heat-Shock Proteins / Q000737:chemistry / Q000378:metabolism*; D007328:Insulin / Q000737:chemistry / Q000378:metabolism; D007546:Isoquinolines / Q000378:metabolism; D007768:Lactalbumin / Q000737:chemistry / Q000378:metabolism; D017017:Methanococcus / Q000378:metabolism*; D018832:Molecular Chaperones / Q000737:chemistry / Q000378:metabolism*; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism; D013696:Temperature; D038201:alpha-Crystallins / Q000737:chemistry / Q000378:metabolism
D019843:Archaeal Proteins; C113795:HSP16.5 protein, Methanococcus jannaschii; D006360:Heat-Shock Proteins; D007328:Insulin; D007546:Isoquinolines; D018832:Molecular Chaperones; D021122:Protein Subunits; D038201:alpha-Crystallins; C093702:lucifer yellow iodoacetamide; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D007768:Lactalbumin; D004229:Dithiothreitol