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Zinc finger arrays have been established as a critical structural feature of proteins involved in DNA recognition. Retroviral nudeocapsid proteins, which are involved in the binding of viral RNA, contain conserved cysteine-rich arrays that have been suggested to coordinate zinc. We provide metalloprotein structural data from an intact virus preparation that validate this hypothesis. Extended x-ray absorption fine structure (EXAFS) spectroscopy of well-characterized and active preparations of equine infectious anemia virus, compared with a peptide with known coordination and in combination with available biochemical and genetic data, defines a Cys3Hisj coordination environment for zinc. The average ofthe Zn-S distances is 2.30(1) A and that of the Zn-N distance (to histidine) is 2.01(3) A.Zinc finger domains are known to be of great importance in a number ofproteins that are involved in nucleic acid binding and transcriptional control. All known retroviral nucleocapsid (NC) proteins contain at least one sequence of the form Cys-Xaa2-Cys-Xaa4-His-Xaa4-Cys, where Xaa represents various amino acids (1-4). Site-directed mutagenesis experiments involving point mutations of cysteine, histidine, and adjacent residues in the NC protein of murine leukemia virus indicate that the conserved arrays are intimately involved in retroviral RNA recognition, since these mutants give rise to virus particles with little or no detectible viral RNA (5, 6). Subsequent genetic experiments with human immunodeficiency virus type 1 (HIV-1) gag domains, which have two cysteine arrays rather than one, reveal that mutations in either Cys-His array reduce RNA packaging considerably, and infectivity even more drastically (7,8).Prior investigations have conclusively shown that synthetic peptides based on these conserved retroviral sequences bind Zn2+ tightly and stoichiometrically (also Co2+ and Cd2+), and a wealth of spectroscopic evidence points to a three-cysteine one-histidine coordination for metals bound to these peptides (4,(9)(10)(11). The relevance of the peptides to virus function has been questioned, since the zinc content of avian myeloblastosis virus is reportedly insufficient to populate the cysteine arrays (12). However, more recent evidence clearly shows that the zinc content of HIV-1 and human T-cell leukemia virus type I (HTLV-I) is present in near stoichiometric amounts to the NC protein cysteine arrays (13). Thus, the potential importance of retroviral cysteine arrays and zinc to the physiological functions of nucleic acid binding and infectivity is illustrated by a wide range of investigations. We describe experiments that probe the zinc environment of concentrated preparations of equine infectious anemia virus (EIAV), a lentivirus (14, 15), using extended x-ray absorption fine structure (EXAFS) spectroscopy. Data obtained from analysis of virus was compared to that from a well-characterized 18-residue retroviral-type zinc finger peptide, based on sequences from HIV-1. This peptide has the sequence Val-Lys-Cys-Phe-As...

Several earlier studies have led to different conclusions about the complex of myosin with MgAMP-PNP. It has been suggested that subfragment 1 of myosin (S1)-MgAMP-PNP forms an S1-MgADP-like state, an intermediate between the myosin S1-MgATP and myosin S1-MgADP states or a mixture of cross-bridge states. We suggest that the different states observed result from the failure to saturate S1 with MgAMP-PNP. At saturating MgAMP-PNP, the interaction of myosin S1 with actin is very similar to that which occurs in the presence of MgATP. 1) At 1 degrees C and 170 mM ionic strength the equatorial x-ray diffraction intensity ratio I11/I10 decreased with an increasing MgAMP-PNP concentration and leveled off by approximately 20 mM MgAMP-PNP. The resulting ratio was the same for MgATP-relaxed fibers. 2) The two dimensional x-ray diffraction patterns from MgATP-relaxed and MgAMP-PNP-relaxed bundles are similar. 3) The affinity of S1-MgAMP-PNP for the actin-tropomyosin-troponin complex in solution in the absence of free calcium is comparable with that of S1-MgATP. 4) In the presence of calcium, I11/I10 decreased toward the relaxed value with increasing MgAMP-PNP, signifying that the affinity between cross-bridge and actin is weakened by MgAMP-PNP. 5) The degree to which the equatorial intensity ratio decreases as the ionic strength increases is similar in MgAMP-PNP and MgATP. Therefore, results from both fiber and solution studies suggest that MgAMP-PNP acts as a non hydrolyzable MgATP analogue for myosin.