In the field of genetic code expansion (GCE), improvements in the efficiency of noncanonical amino acid (ncAA) incorporation have received continuous attention. By analyzing the reported gene sequences of giant virus species, we noticed some sequence differences at the tRNA binding interface. On the basis of the structural and activity differences between Methanococcus jannaschii Tyrosyl-tRNA Synthetase (MjTyrRS) and mimivirus Tyrosyl-tRNA Synthetase (MVTyrRS), we found that the size of the anticodon-recognized loop of MjTyrRS influences its suppression activity regarding triplet and specific quadruplet codons. Therefore, three MjTyrRS mutants with loop minimization were designed. The suppression of wild-type MjTyrRS loop-minimized mutants increased by 1.8-4.3-fold, and the MjTyrRS variants enhanced the activity of the incorporation of ncAAs by 15-150% through loop minimization. In addition, for specific quadruplet codons, the loop minimization of MjTyrRS also improves the suppression efficiency. These results suggest that loop minimization of MjTyrRS may provide a general strategy for the efficient synthesis of ncAAs-containing proteins.

Compared to free-living viruses (< 0.22 m) in the ocean, planktonic viruses in the \"cellular fraction\" (0.22 ~ 3.0 μm) are now far less well understood, and the differences between them remain largely unexplored. Here, we revealed that even in the same seawater samples, the \"cellular fraction\" comprised significantly distinct virus communities from the free virioplankton, with only 13.87% overlap in viral contigs at the species level. Compared to the viral genomes deposited in NCBI RefSeq database, 99% of the assembled viral genomes in the \"cellular fraction\" represented novel genera. Notably, the assembled (near-) complete viral genomes within the \"cellular fraction\" were significantly larger than that in the \"viral fraction,\" and the \"cellular fraction\" contained three times more species of giant viruses or jumbo phages with genomes > 200 kb than the \"viral fraction.\" The longest complete genomes of jumbo phage (~ 252 kb) and giant virus (~ 716 kb) were both detected only in the \"cellular fraction.\" Moreover, a relatively higher proportion of proviruses were predicted within the \"cellular fraction\" than \"viral fraction.\" Besides the substantial divergence in viral community structure, the different fractions also contained their unique viral auxiliary metabolic genes; e.g., those potentially participating in inorganic carbon fixation in deep sea were detected only in the \"cellular-fraction\" viromes. In addition, there was a considerable divergence in the community structure of both \"cellular fraction\" and \"viral fraction\" viromes between the surface and deep-sea habitats, suggesting that they might have similar environmental adaptation properties. The findings deepen our understanding of the complexity of viral community structure and function in the ocean.

The characterization and comparison of microbial sequences, including archaea, bacteria, viruses and fungi, are very important to understand their evolutionary origin and the population relationship. Most methods are limited by the sequence length and lack of generality. The purpose of this study is to propose a general characterization method, and to study the classification and phylogeny of the existing datasets.
We present a new alignment-free method to represent and compare biological sequences. By adding the covariance between each two nucleotides, the new 18-dimensional natural vector successfully describes 24,250 genomic sequences and 95,542 DNA barcode sequences. The new numerical representation is used to study the classification and phylogenetic relationship of microbial sequences.
First, the classification results validate that the six-dimensional covariance vector is necessary to characterize sequences. Then, the 18-dimensional natural vector is further used to conduct the similarity relationship between giant virus and archaea, bacteria, other viruses. The nearest distance calculation results reflect that the giant viruses are closer to bacteria in distribution of four nucleotides. The phylogenetic relationships of the three representative families, Mimiviridae, Pandoraviridae and Marsellieviridae from giant viruses are analyzed. The trees show that ten sequences of Mimiviridae are clustered with Pandoraviridae, and Mimiviridae is closer to the root of the tree than Marsellieviridae. The new developed alignment-free method can be computed very fast, which provides an effective numerical representation for the sequence of microorganisms.