Method rubisco large subunit n-methyltransferase useful for targeting molecules to the active-site vicinity of ribulose-1, 5-bisphosphatet

Abstract

The present invention generally relates to a modified Rubisco large subunit ε N-Methyltransferase (Rubisco LSMT, or RLSMT). The present invention also relates to a modified RLSMT-carbonic anhydrase (RLSMT-CA). This modified RLSMT-CA improves the efficiency of the reduction of CO 2 during photosynthesis, which may increase plant growth rates. The present invention also relates to nucleic acids encoding the modified RLSMT-CA or modified RLSMT. Also, the present invention relates to cells including the modified RLSMT-CA or modified RLSMT, plants containing the modified RLSMT-CA or modified RLSMT, and methods using compositions of the present invention. In addition, the present invention relates to antibodies conjugated to CA which may bind to Rubisco, and antibodies which bind a modified RLSMT-CA. The invention also relates to modified forms of the LS and SS of Rubisco where the modified forms are fusions with CA or biologically active fragments thereof. The present invention provides methods of altering Rubisco carboxylase activity and altering plant growth.

1 . A modified Rubisco large subunit  ε N-methyltransferase (RLSMT), comprising a Rubisco large subunit  ε N-methyltransferase or biologically active fragment thereof linked to a second molecule or biologically active fragment thereof, and wherein the modified RLSMT has the ability to bind a Rubisco.  
   
   
       2 . The modified RLSMT of  claim 1 , wherein the Rubisco large subunit  ε N-methyltransferase is linked to the second molecule by a chemical.  
   
   
       3 . The modified RLSMT of  claim 2 , wherein the chemical is glutaraldehyde.  
   
   
       4 . The modified RLSMT of  claim 2 , wherein the modified RLSMT is a fusion protein, wherein the fusion protein is encoded by a nucleic acid, wherein the nucleic acid comprises a first nucleic acid encoding a Rubisco large subunit  ε N-methyltransferase or biologically active fragment thereof, and wherein the nucleic acid further comprises a second nucleic acid encoding a polypeptide.  
   
   
       5 . The modified RLSMT of  claim 1 , wherein, upon binding of the modified RLSMT to Rubisco, the concentration of the second molecule is increased at the active site of Rubisco.  
   
   
       6 . The modified RLSMT of  claim 1 , wherein, upon binding of the modified RLSMT to Rubisco, the concentration of another molecule is increased at the active site of Rubisco.  
   
   
       7 . The modified RLSMT of  claim 5 , wherein the another molecule is an enzymatic product of the second molecule.  
   
   
       8 . The modified RLSMT of  claim 5 , wherein the second molecule is carbonic anhydrase.  
   
   
       9 . The modified RLSMT of  claim 5 , wherein the second molecule is a phosphatase, 3-phosphoglycerate kinase, glyceraldehydes-3-phosphate dehydrogenase, Rubisco Activase, Phosphoribulokinase, polylysine, or oxygenase.  
   
   
       10 . The modified RLSMT of  claim 6 , wherein the another molecule is CO 2 .  
   
   
       11 . The modified RLSMT of  claim 1 , wherein the second molecule alters Rubisco activity.  
   
   
       12 . A host cell comprising the modified RLSMT of  claim 1 .  
   
   
       13 . The host cell of  claim 12 , wherein the host cell is a plant cell.  
   
   
       14 . The plant cell of  claim 13 , wherein plant growth is altered.  
   
   
       15 . A photosynthetic organism comprising a host cell of  claim 12 .  
   
   
       16 . The modified RLSMT of  claim 1 , wherein the modified RLSMT is a fusion protein, wherein the fusion protein is encoded by a nucleic acid.  
   
   
       17 . An isolated nucleic acid encoding the modified RLSMT of  claim 16 .  
   
   
       18 . An expression vector comprising the nucleic acid of  claim 17  operably linked to an expression control sequence.  
   
   
       19 . A host cell comprising the expression vector of  claim 18 .  
   
   
       20 . A modified Rubisco large subunit  ε N-methyltransferase-carbonic anhydrase (RLSMT-CA), comprising a Rubisco large subunit  ε N-methyltransferase or biologically active fragment thereof linked to a carbonic anhydrase or biologically active fragment thereof, wherein the modified RLSMT-CA has the ability to bind a Rubisco large subunit, and wherein carboxylase activity of Rubisco is altered upon binding of Rubisco to the modified RLSMT-CA.  
   
   
       21 . The modified RLSMT-CA of  claim 20 , wherein the Rubisco large subunit  ε N-methyltransferase is linked to the carbonic anhydrase (CA) by a chemical.  
   
   
       22 . The modified RLSMT-CA of  claim 21 , wherein the chemical is glutaraldehyde  
   
   
       23 . The modified RLSMT-CA of  claim 20 , wherein the modified RLSMT-CA is a fusion protein, wherein the fusion protein is encoded by a nucleic acid, wherein the nucleic acid comprises a first nucleic acid encoding a Rubisco large subunit  ε N-methyltransferase or biologically active fragment thereof, and wherein the nucleic acid further comprises a second nucleic acid encoding a carbonic anhydrase or biologically active fragment thereof.  
   
   
       24 . The modified RLSMT-CA of  claim 20 , wherein the modified RLSMT-CA further comprises an antibody.  
   
   
       25 . The modified RLSMT-CA of  claim 23 , wherein the antibody is linked to a CA or a biologically active fragment thereof.  
   
   
       26 . A host cell comprising the modified RLSMT-CA of  claim 20 .  
   
   
       27 . The host cell of  claim 26 , wherein the host cell is a plant cell.  
   
   
       28 . A plant comprising the plant cell of  claim 27 .  
   
   
       29 . The plant of  claim 28 , wherein growth of the plant is altered.  
   
   
       30 . A photosynthetic organism comprising a host cell of  claim 26 .  
   
   
       31 . An isolated nucleic acid encoding the modified RLSMT-CA of  claim 23 .  
   
   
       32 . An expression vector comprising the nucleic acid of  claim 31  operably linked to an expression control sequence.  
   
   
       33 . A host cell comprising the expression vector of  claim 32 .  
   
   
       34 . An antibody which recognizes and binds to a Rubisco or a Rubisco large subunit, or Rubisco large subunit  ε N-methyltransferase wherein the antibody is linked to a second molecule.  
   
   
       35 . The antibody of  claim 34 , wherein the second molecule is a CA or biologically active fragment thereof.  
   
   
       36 . The antibody of  claim 35 , wherein Rubisco carboxylase activity is altered upon binding of Rubisco to the RLSMT bound to the antibody linked to the CA.  
   
   
       37 . A modified Rubisco subunit-carbonic anhydrase (RSCA), comprising a Rubisco subunit or biologically active fragment thereof linked to a carbonic anhydrase or biologically active fragment thereof, and wherein the modified RSCA has the ability to assemble into Rubisco.  
   
   
       38 . The modified RSCA of  claim 37 , wherein the Rubisco subunit is a Rubisco large subunit or a Rubisco small subunit.  
   
   
       39 . The modified RSCA of  claim 37 , wherein the Rubisco subunit is linked to a carbonic anhydrase by a chemical.  
   
   
       40 . The modified RSCA of  claim 39 , wherein the chemical is glutaraldehyde.  
   
   
       41 . The modified RSCA of  claim 40 , wherein the modified RSCA is a fusion protein, wherein the fusion protein is encoded by a nucleic acid, wherein the nucleic acid comprises a first nucleic acid encoding the Rubisco subunit or biologically active fragment thereof, and wherein the nucleic acid further comprises a second nucleic acid encoding the carbonic anhydrase.  
   
   
       42 . A host cell comprising the modified RSCA of  claim 37 .  
   
   
       43 . The host cell of  claim 42 , wherein the host cell is a plant cell.  
   
   
       44 . The plant cell of  claim 43 , wherein plant growth is altered.  
   
   
       45 . A photosynthetic organism comprising a host cell of  claim 42 .  
   
   
       46 . The modified RSCA of  claim 37 , wherein the modified RSCA is a fusion protein, wherein the fusion protein is encoded by a nucleic acid.  
   
   
       47 . An isolated nucleic acid encoding the modified RSCA of  claim 46 .  
   
   
       48 . An expression vector comprising the nucleic acid of  claim 47  operably linked to an expression control sequence.  
   
   
       49 . A host cell comprising the expression vector of  claim 48 .  
   
   
       50 . A method of altering Rubisco carboxylase activity comprising providing a modified RLSMT-CA, RLSMT, or RSCA, a nucleic acid encoding a RLSMT-CA, RLSMT, or RSCA, or an antibody conjugated to CA.  
   
   
       51 . A method of altering plant growth comprising providing a modified RLSMT-CA, RLSMT, or RSCA, a nucleic acid encoding a RLSMT-CA, RLSMT, or RSCA, or an antibody conjugated to CA.  
   
   
       52 . A nucleic acid comprising SEQ ID No.: 1.  
   
   
       53 . A polypeptide comprising SEQ ID No.: 2.  
   
   
       54 . A vector comprising the nucleic acid of  claim 52 .  
   
   
       55 . A host cell comprising the vector of  claim 54.