HCMV-encoded proteins

HCMV is the human herpesvirus with the largest genome, about 230kb. Two HCMV-encoded proteins, IE-1 and IE-2, have been described to inhibit apoptosis mediated by tumor necrosis factor (TNF)-a, although the mechanism underlying this process has not yet been elucidated (Zhu et al. 1995). It has also been shown that overexpression of IE-2, but not IE-1, can protect smooth muscle cells from apoptosis induced by overexpression of p53 or by doxorubicin (Tanaka et al. 1999). 

More recently, the use of liquid chromatography and tandem MS (LC/MS/MS) has also eased purification and recovery methods. For instance, Vamum and colleagues utilized gel-free two-dimensional capillary LC/MS/MS and Fourier transform ion cyclotron resonance MS to identily and determine the relative abundances of viral and cellular proteins in purified HCMV virions and dense bodies. Analysis of the proteins Irom purified HCMV virion preparations has indicated that the particle contains significantly more viral proteins than previously known. They identified more than 71 HCMV-encoded proteins and 70 host cellular proteins in HCMV virions, which included cellular structural proteins, enzymes, and chaperones [6]. Another study using LC/MS/MS for the adenovirus type 5 pro-teome found a total of 11 protein species from 154 peptides, at a sensitivity of 10 copies per virus and a detection limit of 70 finol for two proteins [36]. 

The protein superfamily of proteases [78, 79], however, is an ideal framework for a directed privileged structure-based masterkey concept. It has already been reported that the 5,5-trans-fused lactam moiety was systematically optimized and explored as a serine protease-directed scaffold by GlaxoSmithKline and has delivered progressible lead compounds for various members of that target class [3], such as thrombin [80, 81], elastase [82, 83], HCMV protease [84, 85], and the hepatitis C virus-encoded NS3-4A protease [86, 87]. Here, the initially identified scaffold was engineered toward the serine protease-wide commonality in substrate binding and processing [3],... 

Following gradient purification of virions, LC/MS/MS was used to identify the components of the HCMV virion [6]. The results were verified by coupling high-accuracy mass measurements with LC and FT-ICR (Fourier transform ion cyclotron resonance) MS. Fifty-nine proteins were identified including 12 proteins encoded by known HCMV ORFs not previously known to reside in virions. The classes of proteins identified included capsid proteins, tegument proteins, glycoproteins, and 12 proteins involved in DNA replication and transcription. Additionally, 12 more viral polypeptides were identified that had not been previously characterized [6]. 

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