PAM1

This research was focused on the isolation and characterization of a PAH-catabolizing mycobacterial strain from the petroleum hydrocarbon-contaminated rhizosphere of alfalfa, as well as on revealing some points of interaction between the microorganism and the plant. Mycolicibacterium sp. PAM1, a pyrene degrader isolated from the niche of interest to us, can catabolize fluoranthene, anthracene, fluorene, and phenanthrene. On the basis of curves of PAM1 growth with different PAHs as the sole carbon sources and on the basis of PAH-degradation rates, we found that pollutant availability to the strain decreased in the sequence phenanthrene > fluorene > fluoranthene ~ pyrene > anthracene. For each PAH, the catabolic products were identified. PAM1 was found to have the functional genes nidA and nidB. New data modeling the 2D and 3D structures, intrinsic structural disorder, and molecular dynamics of the nidA and nidB gene products were obtained. The identified genes and intermediates of pyrene degradation indicate that PAM1 has a PAH catabolic pathway that is peculiar to known mycobacterial pyrene degraders. PAM1 utilized some components of alfalfa root exudates as nutrients and promoted plant growth. The use of mycobacterial partners of alfalfa is attractive for enhancing the phytoremediation of PAH-contaminated soils.

Despite previous structural analyses of bacteriophages, quite little is known about the structures and assembly patterns of cyanophages. Using cryo-EM combined with crystallography, we solve the near-atomic-resolution structure of a freshwater short-tailed cyanophage, Pam1, which comprises a 400-Å-long tail and an icosahedral capsid of 650 Å in diameter. The outer capsid surface is reinforced by trimeric cement proteins with a β-sandwich fold, which structurally resemble the distal motif of Pam1's tailspike, suggesting its potential role in host recognition. At the portal vertex, the dodecameric portal and connected adaptor, followed by a hexameric needle head, form a DNA ejection channel, which is sealed by a trimeric needle. Moreover, we identify a right-handed rifling pattern that might help DNA to revolve along the wall of the ejection channel. Our study reveals the precise assembly pattern of a cyanophage and lays the foundation to support its practical biotechnological and environmental applications.

In the budding yeast Saccharomyces cerevisiae, Svl3 and Pam1 proteins work as functional homologues. Loss of their function causes increased levels of chitin deposition in the cell wall and temperature sensitivity, suggesting their involvement in cell wall formation. We found that the N- and C-termini of these proteins have distinctive and critical functions. They contain an N-terminal part that has a probable 2-dehydropantoate 2-reductase domain. In Svl3, this part can be replaced with the yeast 2-dehydropantoate 2-reductase, Pan5, suggesting that Svl3 and its homologues may be able to mediate 2-dehydropantoate 2-reductase function. On the other hand, Svl3 is recruited to the bud tip and bud neck via multiple localization signals in the C-terminal part. One of such signals is the lysine-rich region located in the C-terminal end. The function and localization of Svl3 are significantly disrupted by the loss of this lysine-rich region; however, its localization is not completely abolished by the mutation because another localization signal enables appropriate transport. Svl3 and Pam1 orthologues are found in cells across fungal species. The Svl3 orthologues of Candida glabrata can complement the loss of Svl3 and Pam1 in S. cerevisiae. C. glabrata cells lacking the SVL3 and PAM1 orthologue genes exhibit phenotypes similar to those observed in svl3∆pam1∆ S. cerevisiae cells. Thus, Svl3 homologues may be generally required for the assembly of the cell wall in fungal cells.