GSJ: Received May 21, 22, 2007: http://wbabin.net/saba/saba78.htm

This invention teaches materials and methods for screening a library of nucleic acids for those which encode an enzyme which covalently modifies a substrate.

Figures 1A, 1B, & 2 delineate the essential features of the nucleic acid libraries used. For simplicity only one nondesirable library member is shown, yet of course many more are anticipated.

As shown in the figures, and of essence to the invention is that every nucleic acid in a library is associated with a substrate of the desired enzyme. In these examples the substrate is identical for each library member nucleic acid.

The encoding nucleic acid may be any form but is preferably dsDNA or mRNA, and the enzyme product is preferably comprises a protein (although others molecules such a RNA are possible). The substrate may be any molecule, but preferably comprises a protein or nucleic acid sequence.

The first step is expression of the encoded products and importantly this must be done under conditions wherein trans modification of substrates is minimized. There are several ways this could be achieved. One is to perform the reactions in micro, nano, or pico wells (1,2). Another is to use micro, nano pico vessels (3,4). Another is to use a display technology (5,6) such that each nucleic acid is associated with its encoded product. Yet another is to use a method such that a product of a support affixed nucleic acid is bound adjacent to its encoding nucleic acid (7,8,9).

Turning to Figure 1A, after expression of the encoded products, only the first cleaves the substrate, and this cleavage dissociates the affinity tag from its adjacent nucleic acid member. Those library members which do not encode the enzyme activity retain their tags and are abstracted from those which have lost their tags.

Figure 1B is a derivation wherein the library members are affixed to a support. Only that nucleic acid encoding an enzyme which cleaves the substrate, is liberated from the support.

Notice the process in Figures 1A & 1B is ideally suited for isolating nucleases such as restriction enzymes, and proteases.

In Figure 2, the modification is covalent addition of an atomic grouping, such as a phosphate, methyl, actyl, or carbohydrate. The corresponding enzymatic activity would be a kinase, methyl transferase, actyl transferase, and glycosyl transferase. Notice that removal of these and other groups from the substrate could also be the enzymatic activity desired.

Subsequent to reaction, the modified substrate and associated enzyme encoding nucleic acid can be isolated, such as via an antibody to the modification. For example an antibody to phosphotyrosine.

If it should be that the above invention is indeed novel any patentable rights I may have, I freely give away.

It is hoped that others will honor the invention as delineated above and by the following claims.

Claims