Human sequencing

Apelins and the Apelin Receptor. Figure 2 Sequence alignment of mammalian and amphibian apelin-36 amino-acid sequences, indicates residues conserved across all the species shown. Residues which differfrom the human sequence are highlighted in red. 

When compared with human monoclonals (half-life 14-21 days), murine monoclonals administered to humans display a relatively short half-life (30-40 h). Chimaerization increased serum half-life by fivefold, with typical values of 230 h being recorded (Table 13.4). A prolonged half-life is desirable if the antibody is to be used therapeutically, as it decreases the required frequency of product administration. Chimaeric antibodies also allow activation of Fc-mediated functions (e.g. activation of complement, etc.), as this domain displays human sequence. 

If in a DNA or protein sequence a single position is variant, this may be called a polymorphism or mutation. The difference is set by convention a mutation is rare, and a polymorphism is common. By implication, a mutation may be deleterious, or in rare cases it may be advantageous for the reproductive fitness of its carriers. A widespread polymorphism, on the other hand, is unlikely to affect the fitness to such an extent. A frequency value of 1 % of the polymorphic allele is usually taken as a threshold between mutation and polymorphism (Ki-mura, 1983 Li, 1997). A polymorphic site is called biallelic if two variants segregate in the population, and multiallelic if there are more than two variants, which is a rare event in human sequences. 

Sequence Repeats. In Section IV, V, and VI, to exclude repetitive elements that might have been inserted recently and would not reflect long-term evolutionary patterns, we used REPEATMASKER [40], leading to a reduction of 40-50% of the human sequence length. 

We have started examining nucleotide compositional strand asymmetries in transcribed regions of human sequences [36]. We have computed the 5ta and 5gc skews [Eq. (1)] for intron sequences since, in contrast to exonic sequences, they can be considered as weakly selected sequences. For each gene, we have concatenated all the introns in a unique sequence (see Section II.A). The distributions of the TA and GC skews, computed on the 14,854 intro-containing... 

Human insulin (emp) insulin of human sequence manufactured by enzymatic modification of porcine insulin... 

Maitland ML, Grimsley C, Kuttab-Boulos H et al. Comparative genomics analysis of human sequence variation in the UGTIA gene cluster. Prmacogenomics J. 2006 6 52-62. 

An endogenous murine gene of choice can also be replaced by human sequences (Stacey et al., 1994). This concept of replacing the endogenous... 

Early attempts to synthesize non-peptide antagonists were hampered by the sequence diversity of the primary sequence of the NhCi receptors from humans and rats, which influenced the potency of the compounds studied (Sachais et al., 1993 Saria, 1999). Appropriate models, with NK1 receptors closer to the human sequence, had to be established in gerbils, rabbits, and guinea pigs. 

Schwartz, B., Ivanov, M.-A., Pitard, B., Escriou, V., Rangara, R., Byk, G. et al. (1999) Synthetic DNA-compacting peptides derived from human sequence enhance cationic lipid-mediated gene transfer in vitro and in vivo. Gene Ther., 6,282-292. 

In a study investigating the allometric relationships of pharmacokinetic parameters for five therapeutic proteins, the allometric equations for clearance and volumes of distribution, however, were found to be different for each protein [102]. This variability was attributed to possible species specificity and immune-mediated clearance mechanisms. Species specificity refers to the inherent differences in structure and activity across species. Minute differences in the amino acid sequence may render an agent inactive when administered to foreign species, and may even generate an immunogenic response. Immunogenicity has been clearly demonstrated in a study with the tumor necrosis factor receptor-immunoglobulin fusion protein lenercept. This all-human sequence protein elicits an immune response in laboratory animals which ultimately results in the rapid clearance of the protein [103]. 

The DNA sequence used by the Human Genome Project was not that of a single person, but a composite from several individuals thus it is a representative human sequence. During the project, the identity of the DNA donors was kept anonymous. More blood samples were collected from volunteers than were actually used, and no names were attached to the samples that were analyzed. Thus, not even the donors knew whether their DNA samples were actually used. 

Urcelay, E., Ward, P., Wiener, S. M., Safer, B. and Kotin, R. M. (1995). Asymmetric replication in vitro from a human sequence element is dependent on adeno-associated virus Rep protein. J. Virol. 69, 2038-2046. 

Mice and humans (and their respective cytokines and receptors) have been evolutionarily separated for over 100 million years. Consequently, the murine interferons differ from all of the human interferons at fifty-six to seventy-two amino-acid positions. Nevertheless, it was possible, simply by recombining human sequences, to produce an interferon with an activity greater than that of the natural mouse protein. Sequences that had been pre-tested in humans to function as a-interferons contained the information necessary to build a protein that functions well with murine cells. Even though it was not possible to reconstruct the exact sequence of murine a-interferon from the human genes, it was possible to reconstruct its function. 

Fig. 2. Selection strategy for antibody humanization that combines CDR grafting with framework fine-tuning. Nonhuman sequences are shown in gray, human sequences are white. The six CDRs of the nonhuman Fab are grafted into a human framework that contains a set of diversified residues. These residues are diversified to allow the selection of either the human or the original nonhuman residue. Reprinted with permission from Rader and Barbas (2000).

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