Enzyme design gets 0 hit

[Anonymous]

Hi,

I’m doing a de novo design of an enzyme to fit in a new molecule. There is a protein structure papers that proposed the interactions between another molecule and residues in the protein and I found two molecules share the same atoms in these interactions. So I transfer these interactions and written them into the cst file. There are a total of 3 blocks. However, I found the program reports 2000+ hits after the first round but 0 hits after the second round. What’s causing this error and what should I do to get more hits?

protocols.match.Matcher: …done

protocols.match.Matcher:  TIMING: Bump grids took 468.42 seconds to compute

protocols.match.upstream.ProteinUpstreamBuilder: Considered 97455393 downstream conformations at residue 52 and found 2767 hits.

protocols.match.downstream.ClassicMatchAlgorithm: Finished round 1 with 2767 hits.

protocols.match.upstream.ProteinUpstreamBuilder: Considered 17129070 downstream conformations at residue 80 and found 0 hits.

protocols.match.downstream.ClassicMatchAlgorithm: Finished round 2 with 0 hits.

protocols.match.downstream.ClassicMatchAlgorithm: Erased 2767 round 1 hits with 0 hits remaining.

apps.public.match.match: Matcher ran for 751 seconds, where finding hits took 751 seconds and processing the matches took 0 seconds.

 

The cst file looks like this:

CST::BEGIN

TEMPLATE:: ATOM_MAP: 1 atom_name: O6 P1 O4

TEMPLATE:: ATOM_MAP: 1 residue3: ANA

TEMPLATE:: ATOM_MAP: 2 atom_name: CD2 NE2 CE1

TEMPLATE:: ATOM_MAP: 2 residue3: HIS

CONSTRAINT:: distanceAB: 3.70 0.30 180.00 0 0

CONSTRAINT:: angle_A: 105.6 20.00 100.00 360.00 1

CONSTRAINT:: angle_B: 64.9 20.00 100.00 360.00 1

CONSTRAINT:: torsion_A: -25.4 10.00 50.00 360.00 1

CONSTRAINT:: torsion_B: -119.3 15.00 0.00 360.00 3

CONSTRAINT:: torsion_AB: -172.9 75.00 5.00 360.00 1



ALGORITHM_INFO:: match

  CHI_STRATEGY:: CHI 1 EX_FOUR_HALF_STEP_STDDEVS

  CHI_STRATEGY:: CHI 2 EX_ONE_STDDEV

      IGNORE_UPSTREAM_PROTON_CHI

ALGORITHM_INFO::END

CST::END



CST::BEGIN

TEMPLATE:: ATOM_MAP: 1 atom_name: N1 C1 C3

TEMPLATE:: ATOM_MAP: 1 residue3: ANA

TEMPLATE:: ATOM_MAP: 2 atom_name: OD2 CG OD1

TEMPLATE:: ATOM_MAP: 2 residue3: ASP

CONSTRAINT:: distanceAB: 2.50 0.30 180.00 0 0

CONSTRAINT:: angle_A: 116.9 20.00 100.00 360.00 1

CONSTRAINT:: angle_B: 126.1 20.00 100.00 360.00 1

CONSTRAINT:: torsion_A: 85.3 10.00 50.00 360.00 1

CONSTRAINT:: torsion_B: 69.7 15.00 0.00 360.00 3

CONSTRAINT:: torsion_AB: -167.8 75.00 5.00 360.00 1



ALGORITHM_INFO:: match

  CHI_STRATEGY:: CHI 1 EX_FOUR_HALF_STEP_STDDEVS

  CHI_STRATEGY:: CHI 2 EX_ONE_STDDEV

      IGNORE_UPSTREAM_PROTON_CHI

ALGORITHM_INFO::END

CST::END



CST::BEGIN

TEMPLATE:: ATOM_MAP: 1 atom_name: O3 P1 O2

TEMPLATE:: ATOM_MAP: 1 residue3: ANA

TEMPLATE:: ATOM_MAP: 2 atom_name: OE1 CD OE2

TEMPLATE:: ATOM_MAP: 2 residue3: GLU

CONSTRAINT:: distanceAB: 5.60 0.30 180.00 0 0

CONSTRAINT:: angle_A: 132.2 20.00 100.00 360.00 1

CONSTRAINT:: angle_B: 81.0 20.00 100.00 360.00 1

CONSTRAINT:: torsion_A: 69.8 10.00 50.00 360.00 1

CONSTRAINT:: torsion_B: 17.8 15.00 0.00 360.00 3

CONSTRAINT:: torsion_AB: 11.0 75.00 5.00 360.00 1



ALGORITHM_INFO:: match

  CHI_STRATEGY:: CHI 1 EX_FOUR_HALF_STEP_STDDEVS

  CHI_STRATEGY:: CHI 2 EX_ONE_STDDEV

      IGNORE_UPSTREAM_PROTON_CHI

ALGORITHM_INFO::END

CST::END

And the pos file is below:

N_CST 3

1: 52

2: 80

3: 532

 

The command I use is:

rosetta_bin_linux_2021.16.61629_bundle/main/source/bin/match.linuxgccdebug @gen_flag -match:scaffold_active_site_residues_for_geomcsts design.pos -match:geometric_constraint_file design.cst

While the gen_flag file includes several options:

-s protein_monomer_apo.pdb

-match:lig_name ANA

-extra_res_fa ANA.params

-use_input_sc

-match:output_format CloudPDB

-match_grouper SameSequenceAndDSPositionGrouper

-match:bump_tolerance 0.4

-output_matchres_only false

 

Many thanks for any replies in advance!

 

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