Patent protection of gene mutants in China
date: 2022-06-17 Tianyun Jin Read by:

With the development of biotechnology, a variety of enzymes have been used in industrial production, such as enzyme-catalyzed synthesis of compounds, especially chiral drugs or chiral drug intermediates. Compared with the traditional chemical synthesis that may require a large amount of organic solvent, toxic reagents or expensive metal catalysts, enzyme-catalyzed synthesis of compounds avoids the use of a large amount of organic solvent, and has the advantages of mild reaction conditions and environmental friendliness, etc. Some enzymes can also efficiently catalyze the production of chiral compounds and avoid the separation of chiral compounds. Therefore, it has attracted more and more attention of enterprises and researchers. However, most wild-type enzymes have defects such as narrow substrate range, low activity and poor stability, which are still far from industrial application. In order to make the enzymes applied to industrial production, researchers usually need to modify the enzymes by genetic engineering, so as to obtain the enzyme mutations with optimized performances.

At present, the genetic modification of enzymes usually includes gene mutation, such as site-directed mutation. In order to obtain an enzyme with a certain performance, researchers usually mutate the coding gene of the enzyme first, so as to construct a batch of mutants with different mutation sites, and then screen the performances of these mutant enzymes. Some mutation sites that can improve the performance of the enzyme were further combined to obtain mutants with further improved performance; or on this basis, combined with computer software to simulate the structure of the enzyme, further select sites for site-directed mutation, saturation mutation or directed evolution, and constantly improve the performances of the enzyme.

When the mutants with optimized performance are obtained, enterprises usually apply for patents in order to protect intellectual property rights. So, what problems will be encountered in the patent application for such gene mutants in China? Next, discuss with examples (this case is fictitious).Researchers of a company first mutated at ransaminase with SEQ ID No: 1 through site mutation, and then through experimental verification, it was found that the transaminase mutants with mutation site of G17VL36PQ40HG69YH70TL73Aor V77G have been greatly improved in enzyme activity. Then on this basis, the combined mutants of the above mutation sites were constructed respectively. It was found that the activity of the mutations with mutation site combinations of G17V+L36P+Q40HG69Y+H70TL36P+L73A+V77GQ40H+G69Yor L 36P+Q40H+G69Y was further improved on the basis of mutation with single mutation site. Since then, in order to further improve the tolerance to organic solvents of the transaminase mutants, the researchers carried out a series of site-directed mutations on the basis of these combined mutations, and obtained a large number of mutants with greatly improved transaminase activity and organic solvent tolerance. In view of this, in order to obtain the widest possible protection, the applicant has written the following claims:


1.A transaminase mutant, wherein an amino acid sequence of the transaminase mutant is an amino acid sequence obtained by mutation occurred in an amino acid sequence as shown in SEQ ID NO: 1, and the mutation at least comprises one of the following mutation sites: G17VL36PQ40HG69YH70TL73Aor V77G,or the transaminase mutant has the mutation sites of the mutation, has more than 80% identity with the amino acid sequence of the mutation, and has transaminase activity.

2. The transaminase mutant according to claim 1, where in the mutation comprises any one of the following mutation site combinations: g17v + l36p + q40h, g69y + h70t, l36p + l73a + v77g, q40h + g69y or l 36p + q40h + g69y.


Such claims include mutants with single point mutations and combination mutations. The limitation of "homology" can avoid competitors from using it with a little modification on the basis of these mutants without tort. However, can such claims be patented? From the current judicial practice, the answer is No.


As we all know, to be granted, a patent application must be novel, creative and practical. In addition, the claims must have a single general inventive concept, possess the unity and be support by the specification.

For this case, first of all, there is no problem with practicability. It is assumed that SEQ ID No: 1 is a known sequence, but the mutation sites have not been disclosed. Then, it is novel. As for creativity, it depends on the above mutations of SEQ ID No: 1, which has greatly improved the enzyme activity, which is unexpected for those skilled in the art in the field of biotechnology with low predictability of technical effect. Therefore, based on unexpected technical effects, the enzyme mutants are creative. Then the problem comes. In this case, each mutation site becomes a "specific technical feature". According to paragraph 1 of Article 31 of the patent law, two or more inventions or utility models belonging to a general inventive concept may be filed as one application. Two or more inventions or utility models belonging to a general inventive concept which may be filed as one application shall be technically inter-related and contain one or more of the same or corresponding special technical features. The expression "special technical features" shall mean those technical features that define a contribution which each of those inventions or utility models, considered as a whole, makes over the prior art. Therefore, the single point mutations do not belong to a general inventive concept. To be protected in one patent application, the enzyme mutants must have at least one "specific technical feature", that is, they contain at least one of the same mutation sites. Based on this situation, will the R & D strategy of researchers be adjusted appropriately in order to better protect the later R & D results at reasonable cost?

Secondly, as per the limitation of "homology", quote the examiner’s comments of similar patent as "For the mutants defined by "homology", although the formed sequence has certain sequence structure similarity with the original sequence, of course, there are certain differences of sequence structure, and the differences of sequence structure may change the three-dimensional structure of the enzyme, resulting in the change or even loss of its function. Moreover, the specification of this application does not provide an example of amino acid sequence meeting the above conditions and verify its technology effect. Therefore, those skilled in the art cannot expect that all amino acid sequences defined by homology can solve the technical problem to be solved in the application according to the contents fully disclosed in the specification and the prior art. The mutants with open restriction include a large number of mutation sites and different combinations of the sites. The above mutations contain a large number of transaminase mutants whose functions have not been verified, while the specification of this application only verifies the transaminase activity of single point mutants at some sites, some double site and three site combination mutants. Those skilled in the art cannot expect which mutants and their combinations in the above mutations can change the spatial conformation of the protein and affect its function. Moreover, the application does not describe the spatial conformation, functional domain and other information of the enzyme. Therefore, those skilled in the art cannot expect that other mutants other than those verified in the specification of the application still have the function of transaminase and can solve the technical problems to be solved in the application. To sum up, the mutant defined in claim 1 contains the content speculated by the applicant, and its effect is difficult to determine in advance. Therefore, claim 1 is not supported by the description ".

In this case, is the scope of protection finally only the mutant verified by the embodiment in the specification? In fact, it is not absolute. It also depends on the specific contents disclosed in the specification. For example, the five patent invalidation cases filed by Yichang East Sunshine Pharmaceutical Co., Ltd. for the "glucoamylase" series patents owned by Dennisco, the protection of amino acids / nucleotides with mutation sites is involved, and the judgment of the Patent Reexamination Board of the State Intellectual Property Office of which has certain guiding significance. Herein, for CN200880116695.2, the view of the collegiate panel is "When some single point mutations and multiple different combination mutations of these single point mutations have improved performance, it can be reasonably speculated that these single point mutations or combination mutations are related to this performance. Since there are such single point mutations related to performance in the mutation combinations in claims 1, 2 and 3, based on this, the mutation combinations in claims 1, 2 and 3 may also have the same or similar improved performance. The newly combined mutants in claims 1, 2 and 3 can be supported by single mutants and other combined mutants ". Therefore, is it possible to protect a series of combinatorial mutants involving these single point mutations in an open restriction mean as long as it is proved that the single point mutants and several related combinatorial mutations have good performance? As we know, if the specification discloses the relevant research on the spatial conformation and functional domain of the enzyme, the applicant can summarize a reasonable scope based on this, which is "larger range than the mutants verified in the embodiment". However, the current situation is that the research and development of many enterprises are for practical application. Therefore, in the process of mutant research and development, they only need to obtain mutants suitable for industrial production according to their final performance, and there is no need to spend too much manpower and material cost to study the spatial conformation and functional domain of enzyme. In this case, the above cases can give some guidance.

The predictability of the technical effect in the biological field is low, and the experiments can not exhaust all mutation schemes. How to summarize the reasonable scope according to the existing embodiments, or provide what kind of data can support the scope you want to protect, according to the legislative purpose of "matching the protection scope with the technical contribution", it is worthy of serious consideration by the applicant, patent agency and patent examination agency.