Agnoprotein is an important regulatory protein of the human polyoma JC virus (JCV) and plays critical roles during the viral replication cycle. It forms highly stable dimers and oligomers through its Leu/Ile/Phe-rich domain, which is important for the stability and function of the protein. We recently resolved the partial 3D structure of this protein by NMR using a synthetic peptide encompassing amino acids Thr17 to Gln52, where the Leu/Ile/Phe- rich domain was found to adopt a major alpha-helix conformation spanning amino acids 23 to 39. Here, we report the resolution of the 3D structure of full-length JCV agnoprotein by NMR, which not only confirmed the existence of the previously reported major α-helix domain at the same position but also revealed the presence of an additional minor α-helix region spanning amino acid residues Leu6 to Ala10. The remaining regions of the protein adopt an intrinsically unstructured conformation.

Both Pfizer-BNT162b2 and Moderna-mRNA-1273 vaccines can elicit an effective immune response against SARS-CoV-2 infection. However, the elicited serum antibody levels vary substantially and longitudinally decrease after vaccination. We examined the correlation of vaccination-induced IgG levels and neutralization titers against newly emerged variants remains and demonstrate a significant reduction of neutralization activities against the variants (B.1.1.7, B.1.525, and B.1.351) in Pfizer or Moderna vaccined sera. There was a significant and positive correlation between serum IgG levels and ID50 titers for not only SARS-CoV-2 WT but also the variants. These findings indicate that a high level of anti-spike IgG may offer better protection against infection from SARS-CoV-2 and its variants. Therefore, it is necessary to longitudinally monitor specific serum IgG level for evaluating the protective efficacy of the vaccines against SARS-CoV-2 and its new variants.

The ubihydroquinone-cytochrome c oxidoreductase (or the cytochrome bc 1 complex) fromRhodobacter capsulatus is composed of the Fe-S protein, cytochrome b, and cytochromec 1 subunits encoded bypetA(fbcF), petB(fbcB), and petC(fbcC) genes organized as an operon. In the work reported here, petB(fbcB) was split genetically into two cistrons, petB6 andpetBIV, which encoded two polypeptides corresponding to the four amino-terminal and four carboxyl-terminal transmembrane helices of cytochrome b, respectively. These polypeptides resembled the cytochromeb 6 and su IV subunits of chloroplast cytochromeb 6 f complexes, and together with the unmodified subunits of the cytochromebc 1 complex, they formed a novel enzyme, named cytochrome b 6 c 1complex. This membrane-bound multisubunit complex was functional, and despite its smaller amount, it was able to support the photosynthetic growth of R. capsulatus. Upon further mutagenesis, a mutant overproducing it, due to a C-to-T transition at the second base of the second codon of petBIV, was obtained. Biochemical analyses, including electron paramagnetic spectroscopy, with this mutant revealed that the properties of the cytochromeb 6 c 1 complex were similar to those of the cytochrome bc 1 complex. In particular, it was highly sensitive to inhibitors of the cytochrome bc 1 complex, including antimycin A, and the redox properties of its b- and c-type heme prosthetic groups were unchanged. However, the optical absorption spectrum of its cytochrome b L heme was modified in a way reminiscent of that of a cytochromeb 6 f complex. Based on the work described here and that with Rhodobacter sphaeroides (R. Kuras, M. Guergova-Kuras, and A. R. Crofts, Biochemistry 37:16280–16288, 1998), it appears that neither the inhibitor resistance nor the redox potential differences observed between the bacterial (or mitochondrial) cytochromebc 1 complexes and the chloroplast cytochromeb 6 f complexes are direct consequences of splitting cytochrome b into two separate polypeptides. The overall findings also illustrate the possible evolutionary relationships among various cytochrome bcoxidoreductases.