title: Pea lectin unfolding reveals a unique molten globule fragment chain.
abstract: Pea lectin (PSL) is a dimeric protein in which each subunit comprises two intertwined, post-translationally processed polypeptide chains--a long β-fragment and a short α-fragment. Using guanidine hydrochloride-induced denaturation, we have investigated and characterized the species obtained in the unfolding equilibrium of PSL by steady-state and time-resolved fluorescence, phosphorescence, and selective chemical modification. During unfolding, the fragment chains become separated, and the unfolding pattern reveals a β-fragment as intermediate that has the molten globule characteristics. As examined by 8-anilino-1-naphthalenesulfonate (ANS) binding, the fragment intermediate shows ~20 fold increase in ANS fluorescence, and a large increase in ANS lifetime (12.8 ns). The tryptophan environment of the molten globule β-fragment has been probed by selective modification with N-bromosuccinimide (NBS), which shows that two tryptophans, possibly Trp 53 and Trp 152 are oxidized while the other Trp 128 remains resistant to oxidation. The different types of tryptophan environment for the intermediate are supported by phosphorescence studies at 77 K, which gives a (0,0) band at 410 nm. These results seem to indicate that the larger fragment chain of PSL can independently behave as a monomeric or single domain protein that undergoes unfolding through intermediate state(s), and may provide important insight into the folding problem of oligomeric proteins in general and lectins in particular.
authors.affiliation: Department of Chemistry & Biochemistry, Presidency University, 86/1 College Street, Kolkata, India.
authors.forename: Debasish
authors.initials: D
authors.lastname: Sen
authors.forename: Dipak K
authors.initials: DK
authors.lastname: Mandal
mesh_terms: D020106:Acrylamide / Q000737:chemistry; D001974:Bromosuccinimide / Q000378:metabolism; D019791:Guanidine / Q000494:pharmacology; D008958:Models, Molecular; D009282:Naphthalenesulfonates / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry* / Q000378:metabolism; D037121:Plant Lectins / Q000737:chemistry* / Q000378:metabolism; D017510:Protein Folding / Q000187:drug effects; D017433:Protein Structure, Secondary; D058767:Protein Unfolding / Q000187:drug effects; D013050:Spectrometry, Fluorescence; D013696:Temperature
chemical_list: D009282:Naphthalenesulfonates; D010446:Peptide Fragments; D037121:Plant Lectins; C036292:pea lectin; C435043:1-naphthalenesulfonic acid; D020106:Acrylamide; D019791:Guanidine; D001974:Bromosuccinimide