Could protein tertiary structure influence mammary transgene expression more than tissue specific codon usage?

Abstract

Animal mammary glands have been successfully employed to produce therapeutic recombinant human proteins. However, considerable variation in animal mammary transgene expression efficiency has been reported. We now consider whether aspects of codon usage and/or protein tertiary structure underlie this variation in mammary transgene expression.

Fig. 1

A dendogram reflecting the codon usage of 22 genes selectively expressed in heart (red) and 17 genes selectively expressed in human kidney (black). The pairwise distances underlying this tree reflect the degree to which the genes differ in their codon usage. As this tree demonstrates, heart-specific genes can generally be distinguished from heart-speicific genes purely on the basis of their synonymous codon usage. The observed separation between these two classes of genes would not have occurred by random chance (P < 0.001)

Fig. 2

Expression levels of recombinant human proteins in the milk of transgenic animals. The expression level of each protein presents the highest one among different studies summarized in Table 1; ATCD20 IgH and ATCD20 IgL indicate the heavy and light chain of human anti-CD20 monoclonal antibody; FGA, FGB and FGG note the alpha chain, beta chain and gamma chain of fibrinogen (FIB) respectively; mCol18a1 stands for mouse collagen, type XVIII, alpha 1; star notes the expression level of recombinant protein with a cDNA based expression construct; open circle indicates the expression level values of hFVIII(0.0027 mg/mL), hIL-2(0.000043 mg/mL) and mCol18a1(0.00003 mg/mL) which all are too low to be fully presented on the top of the bar

Fig. 3

Dendograms reflecting the codon usage of milk proteins of the main five livestock (red) and recombinant proteins. The top 13 high level expressed recombinant proteins (≧5 mg/mL) are showed in blue, and moderate and lower expressed recombinant proteins are indicated in black. The three classes of genes (indicated in three different colour) can not be clearly discriminated from each other (cow, P = 0.992; goat, P = 0.982; sheep, P = 0.992; rabbit, P = 0.962; pig, P = 0.994)

Fig. 4

The tertiary structures of the main domains of casein proteins, 10 recombinant human proteins highly expressed in the milk of transgenic animal, and two moderately expressed proteins CEL (1.00 mg/mL) and Calc1(2.10 mg/mL) based on cDNA expression constructs. Figures were made using VMD (http://www.ks.uiuc.edu/Research/vmd/) and rendered using Snapshot