Mutations of the coat protein gene of bacteriophage λ that overcome the necessity for the FI gene; the EFi domain

Abstract

SummaryThe functions of most of the 10 genes involved in phage capsid morphogenesis are well understood. The function of the FI gene is one of the exceptions. Mutants in FI fail to mature and package DNA. The gene product (gpFI) seems to act as a catalyst for the formation of an intermediate in capsid assembly called complex II, which contains a procapsid (an empty capsid precursor), terminase (the enzyme that cleaves the DNA precursor and packages it into the procapsid) and DNA. The mechanism for this stimulation remains unknown. It has also been reported that gpFI appeared to stimulate terminase-mediated cos cleavage, in the absence of procapsids, by increasing enzyme turnover. In comparison with other head-gene mutants, FI mutants are leaky, producing approx. 0.1 phage per infected cell. Some second-site revertants of FI ¹ phages, called 'fin', that bypass the necessity for gpFI, have been isolated and found to harbour a mutation in the genes that code for the two subunits of terminase. In the course of mapping additional fin mutants, it was discovered that some mapped outside the terminase genes. To localize the mutations, restriction fragments of fin mutant DNAs were subcloned into plasmids and their ability to contribute to fin function was determined by marker-rescue analysis. The location of the fin mutation was further delineated by deletion analysis of a plasmid that was positive for fin. This showed that some fin mutations mapped to a region comprising genes E, D and a portion of C. The sequencing of this entire region in several fin isolates showed that the fin mutations are clustered in a small region of gene E corresponding to a portion of 26 amino acid residues of the coat protein (gpE ). We have called this region of the protein the EFi domain.All the mutations result in an increase in positive charge relative to the wild-type protein. These results suggest that DNA maturation and packaging are in part controlled by an interaction between gpFI and capsid gpE.

Microbiological and molecular impacts of AbiK on the lytic cycle of Lactococcus lactis phages of the 936 and P335 species

“…In phage lambda, mutations affecting late genes involved in head morphogenesis can provoke similar accumulation of concatenated DNA (Catalano et al, 1995 ;Murialdo, 1991 ;Murialdo & Becker, 1978 ;Murialdo & Tzamtzis, 1997). Recently, a similar organization of the late genes was observed between lambda and the lactococcal phage sk1, a close relative of P008 and p2 (Chandry et al, 1997).…”

Structural and Biochemical Characterization of Phage λ FI Protein (gpFI) Reveals a Novel Mechanism of DNA Packaging Chaperone Activity

“…This number is much higher than that observed for amber mutants of completely essential genes like gpE that produce fewer than 10 Ϫ6 particles/ cell (10). Mutants of that are able to grow in the absence of gpFI have been isolated and produce small plaques, suggesting a partial rescue of activity due to inefficient packaging (9,(11)(12)(13). These mutations, termed fin (FI-independent), map to the terminase and major head genes.…”

“…finA mutants, which induce a 4-fold increase in gpA expression and result in a 10-fold increase in terminase activity, map to the terminase genes Nu1 and A. finB mutants do not affect the expression of gpA and constitute missense mutations in Nu1 and E. The effects of the Nu1 finB mutants are unknown, but it is speculated that they increase terminase cleavage activity (12). The finB mutations that map to gene E cluster in a region encoding a 26-amino acid stretch termed the EFi domain (11). Each of the mutations involves a single amino acid substitution that results in a net increase in positive charge on the capsid and has a phenotype only in the absence of FI.…”

Efficient DNA Packaging of Bacteriophage PRD1 Requires the Unique Vertex Protein P6

“…However, there are certain similarities between P6 and bacteriophage lambda packaging factor gpFI, which is involved in mediating terminase binding to the lambda procapsid and the initial cos cleavage of the DNA (60,61,86). Similar to what is seen with gpFI in bacteriophage lambda, lack of PRD1 P6 decreases but does not totally inhibit packaging, and the binding of packaging ATPase P9 to the capsid is altered although, obviously, no cos-type cleavage is necessary for the unit-length PRD1 genome.…”

“…Some of these proteins are probably part of the actual packaging machinery. For example, lambda gpFI may act by promoting binding of the DNA-terminase complex to the prohead (13,60,61). Other proteins may act by sealing or "plugging" the packaged capsids and preventing release of the packaged DNA; examples include the head completion proteins gp15 and gp16 that bind to the portal of DNA-filled SPP1 virions (14,54,70), bacteriophage lambda gpW and gpFII (62,74) and, possibly, the bacteriophage P22 head completion proteins gp4, gp10, and gp26 (87).…”