Identification and Crystallisation of a Heat- and Protease-Stable Fragment of the Bacteriophage T4 Short Tail Fibre
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Mark J. van Raaij
Abstract
Irreversible binding of Teven bacteriophages to Escherichia coli is mediated by the short tail fibres, which serve as inextensible stays during DNA injection. Short tail fibres are exceptionally stable elongated trimers of gene product 12 (gp12), a 56 kDa protein. The Nterminal region of gp12 is important for phage attachment, the central region forms a long shaft, while a Cterminal globular region is implicated in binding to the bacterial lipopolysaccharide core. When gp12 was treated with stoichiometric amounts of trypsin or chymotrypsin at 37 C, an Nterminally shortened fragment of 52 kDa resulted. If the protein was incubated at 56 C before trypsin treatment at 37 C, we obtained a stable trimeric fragment of 3 33 kDa lacking residues from both the N and Ctermini. Apparently, the protein unfolds partially at 56 C, thereby exposing proteasesensitive sites in the Cterminal region and extra sites in the Nterminal region. Welldiffracting crystals of this fragment could be grown. Our results indicate that gp12 carries a stable central region, consisting of the Cterminal part of the shaft and the attached Nterminal half of the globular region. Implications for structure determination of the gp12 protein and its folding are discussed.
Copyright © 2001 by Walter de Gruyter GmbH & Co. KG
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