Abstract:
:We report and interpret the first Raman spectrum of a double-stranded RNA virus containing a membrane envelope. Spectra of the native bacteriophage phi 6 and of its isolated host-attachment (spike) protein and phospholipid-free core assembly were collected from aqueous solutions over a wide range of temperature. Comparison of the vibrational spectra by digital difference methods permits the following structural conclusions regarding molecular constituents of the fully assembled virion. (1) The double-stranded RNA, phospholipid and protein components of the phage exhibit Raman amplitudes in accordance with their biochemically determined compositions in the native virion (10, 20 and 70%, respectively). (2) alpha-Helix and irregular conformations are the dominant secondary structures in proteins of both the viral membrane and nucleocapsid. This represents a departure from previously examined icosahedral phage and plant viruses, which are dominated by beta-sheet structures. (3) The phospholipids of the viral membrane are liquid crystalline throughout the determined range of virus thermostability (0 to 40 degrees C). (4) The P3 spike protein of phi 6, which is anchored to, but not sequestered within the viral membrane, is largely alpha-helical (approximately 35%) and highly thermolabile. Denaturation of P3 at temperatures above 30 degrees C leads to appreciable loss (approximately 20%) of alpha-helix in favor of beta-strand structure, and alters significantly the environments of many aromatic side-chains. (5) The secondary structures of integral membrane proteins of phi 6 are overwhelmingly alpha-helical (approximately 70 to 80%) and also thermolabile. In contrast to P3, which exhibits aspartate and glutamate carboxyls in the ionized form (CO2-), the integral membrane proteins exhibit only protonated carboxyl groups (COOH). Treatment of phi 6 with butylated hydroxytoluene (BHT), which has been shown to remove the P3 spike protein, does not significantly perturb phospholipids and associated integral proteins of the viral membrane or structural proteins and packaged double-stranded RNA of the nucleocapsid. However, P3 subunits, which are recovered after BHT treatment, exhibit radically altered secondary and tertiary structures, including the loss of most subunit alpha-helices. Among the P3 side-chains affected by BHT treatment, we note a general trend toward greater hydrophilicity and greater solvent exposure of the aromatic residues Trp and Tyr. On the other hand, the cysteine sulfhydryl groups of the BHT-isolated P3 monomer are not solvent exposed and function as strong hydrogen-bond donors in the protein core.(ABSTRACT TRUNCATED AT 400 WORDS)
journal_name
J Mol Bioljournal_title
Journal of molecular biologyauthors
Li T,Bamford DH,Bamford JK,Thomas GJ Jrdoi
10.1006/jmbi.1993.1163subject
Has Abstractpub_date
1993-03-20 00:00:00pages
461-72issue
2eissn
0022-2836issn
1089-8638pii
S0022283683711630journal_volume
230pub_type
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