PMC

Interactions among functional groups of proteins. Each row and column of the shown profile corresponds to a protein-protein interaction (two-hybrid) count with different functional classes (see matrix). The interactions within certain functional classes are enriched compared to other functions groups, e.g. head assembly proteins show 15 interactions among each other, 8 interactions are detected between tail assembly proteins and 3 interactions among proteins of unknown function (see Additional file : Tables S4 and S5 for details).

The protein interaction network of phage lambda. Interactions from this study have been integrated with previously published interactions ("literature"). Nodes in the network represent proteins and are colored according to their functional class (see color key). The protein-protein interactions are indicated by lines ("edges"). The edge color represents the source of the interactions, e.g., all red edges are previously reported interactions, all blue interactions were identified in our two-hybrid study, and all green interactions are previously known and are reproduced in our study.

The Lambda genome and virion. (A) Genome of phage lambda. Colored ORFs correspond to colored proteins in (B). Main transcripts are shown as arrows. (B) A model of phage lambda, indicating protein-protein interactions. Proteins in bold font have known structures (Table 1). Numbers indicate the number of protein copies in the particle. It is unclear whether M and L proteins are in the final particle or only required for assembly. (C) Electron micrograph of phage lambda. (A) and (C) modified after [].

Tail assembly. The lambda tail is made of at least 6 proteins (U, V, J, H, Tfa, Stf) with another 7 required for assembly (I, M, L, K, G/T, Z). Assembly starts with protein J, which then, in a poorly characterized fashion, recruits proteins I, L, K, and G/T to add the tape measure protein H. G and G/T then leave the complex so that the main tail protein (V) can assemble on the J/H scaffold. Finally, U is added to the head-proximal end of the tail. Protein Z is required to connect the tail to the pre-assembled head. Protein H is cleaved between the action of U and Z []. It remains unclear if proteins M and L are part of the final particle []. Modified after [].

Head assembly. Head assembly has been subdivided in five steps although most steps are not very well understood in mechanistic terms. The tail is assembled independently. The C protease, the scaffolding protein Nu3, and the portal protein (B) form an ill-defined initiator structure. Protein E joins this complex but the chaperonins GroES and GroEL are required for that step. Within the prohead C and E are processed to form covalently joined X1 and X2 proteins although this is controversial (see text). Proteins Nu1, A, and FI are required for DNA packaging. Protein D joins and stabilizes the capsid as a structural protein. FII and W are connecting the head to the tail that joins once the head is completed. Modified after [] and [].

Yeast two-hybrid array screens and vectors. Shown are two Y2H screens with four different vector combinations. Each interaction is represented by two colonies to ensure reproducibility. (A) Lambda bait protein A (DNA packaging protein) was fused to an N-terminal DNA-binding domain ("DBD", in pGBKT7g) and was tested against prey constructs in both N- and C-terminal configurations (activation domains in pGADT7g, and pGADCg). (B) The C-terminal DBD fusion (in pGBKCg) as tested against prey constructs in both N- and C-terminal configurations (in pGADT7g, and pGADCg). The interactions of C-terminal preys are labeled with an asterisk (*), all remaining interactions use N-terminal fusions. All the interactions obtained from the array screening were subjected to Y2H retests: we were able to retest all the interactions shown in Figure 2 except A-Ea47, which has thus been removed from the final interaction list. Technical details of the screening procedure have been described in [,]. (C) Interaction quality assesment. Using the experimental derived false positive rate from [] and Bayes theorem, we estimated the probability of an interaction to be true. This estimate depends on the vector system, being highest (83%) for pDEST22/32, and lowest (40%) for pGBKCg/pGADT7g. (D) Detection of known PPIs with different vector systems. Known PPIs are enriched in the subset of PPIs detected by > = 2 vector systems compared to PPIs detected by 1 vector combination.