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Supplementary Table 1: Surface analysis by subfamily annotation of organic solvent stable lipases
Subfamily 1
SNL
Temperature
stability
Mesostable
pH stability
Organic stability
Substrate
Application
LogP
*Surface properties
Alkalophilic
Petroleum spirit, heptane, toluene,
DMF, and DMSO, diisopropylfluorophosphate.
Long chain
Organic
synthesis
Wide
N and Ne residues more SE. Surface
exposure of D > E
Res less Hp till 70-80%. From 6070% till 30-40% more P and H SE.
No C present. W and H buried.
Q> SE. A more buried. L more
buried.
Unstable in high concentration of
PMSF, SDS, acetone, benzene,
ethanol, and methanol
EDTA has no affect.
Subfamily 2
YLL
ANLO44
Temperature
stability
Thermostable
Thermostable
pH stability
Organic stability
Substrate
Application
LogP
*Surface properties
8
Water miscible organic solvents, 10%
acetone, methanol , ethanol,
isopropanol, DMSO. Maximum
activity in DMSO.
Long chain
C12-c16
Biodissel,
organic
synthesis
-1.12-0.9
More Ne and N SE. Hp less SE.
C29 20-30%, C42 20-30% and
C243 10-20% SE. 2 C buried.
M100 50-60% SE. K and H
more SE exposed than R. More
D SE. W SE. G more buried.
Q>SE. 2 DSB between residues
42 -298 and 119-112.
L more buried.
1,3 regiospecific
Middle chain
Biodissel,
organic
synthesis
-0.55 -0.25
> Ne and N SE, Hp and P less
SE. C104 30-40%, C35 1020% SE. 5 cys buried. D>E SE.
> K and H SE than R.. M209
30-40% SE. M132 & M50
burtied. W SE.
Acidicneutral
In 20% organic solvents, there was no
activity left in acetone, ethanol,
isopropanol, 60% activity remaining in
methanol and 95% activity remaining
in DMSO.
Stable in Methanol, 2-propanol &
methyl acetate
No deactivation in presence of 50 %
water-miscible organic solvents.
However, water-immiscible
aliphatic solvents reduce the lipase
1
activity between 20 and 80 %.
ATL
Wide range of
stability
3-12
Stable in various organic solvents
1,3 regiospecific
stable over 410
Inhibited by ionic detergents, whereas
non-ionic detergents stimulates
enzyme activity.
Wide
Complicated exposure of
surface charges. C273 and
C110 20-30% and 109-20% SE
respectively. 4 cys buried. D
and E equally SE. >K res SE.
M1 60-70% SE. H< SE. W SE.
1 DSB between residues 110113.
Organic
synthesis
-1.12- 3.5
Medium chain fatty
acids; hydrophobic,
crevice-like binding
site located near the
protein surface
flexible substrates are
preferably
hydrolyzed in sn-1,
rigid substrates in sn3
Do not bind tertiary
alcohol
Organic
synthesis
Log P >2
More Ne SE till 60-70%. N and
Hp balance each other. >D SE.
> K SE. H buried. M243 6070% SE. M300 40-50% SE.
M292 20-30% SE. 1M buried.
6 C buried. W SE. 1 DSB
between residues 36-41. A, L
buried
P and N more SE. C266 3040% SE. C38 20-30% SE.
M154 buried. No SE M. H res
buried. More K SE. >D SE. 4 C
buried. W more SE. Maximum
Q SE. 1 DSB between residues
27-38.
-doBind tertiary alcohol
Detergent
Organic
synthesis
>4
Unaffected by EDTA or by 2mercaptoethanol and potassium
ferrocyanide
FHL
Thermostable
Acidicneutral
50% DMSO, hexane, benzene.
Middle chain fatty
acid
1,3- regiospecific
ROL
Mesostable
Neutral
Fairly stable in alkanes and long chain
alcohols. Most stable in hexane.
Denatured in hydrophilic solvents such
as acetone or short chain (C1–C3)
alcohol. Most stable in hexane.
CTAB reduces enzyme activity
RML
Mesostable
Alkalophilic
80 mM Tween 80 or 90 mM Tween
20, hexane.
2
N and Hp more SE. Ne more
SE. D>E SE. His SE. W SE.
Y, M buried. 1 DSB between
residues 182-240.
Subfamily 3
Temperature
stability
pH stability
Organic stability
PFL
Thermostable
Alkalophilic
Water-miscible organic solvents solutions
and highly tolerant to some water-immiscible
organic solvents.
Stable in isopropanol, methanol and ethanol,
as well as acetone and glycerol
Inactivated in cyclohexane,
benzene and acetic acid.
ASYL
Thermostable
Alkalophilic
Substrate
Applicatio
n
LogP
*Surface properties
Many
Unable to
predict
R res more SE. C buried. E>D SE. P
and N more SE. W SE. His SE. Q
more buried than SE. Q more buried.
Biodissel
-0.55-3.5
Organic
synthesis
>2
Medium and
the long-acyl
chain esters
funnel-like
substratebinding sites
Biodissel
>0
Ne and Hp more SE. P and N more
buried than SE. C15 and C20, 3040% SE, C214 and C264 SE. E>D
SE. K. M is absent. W buried. H
buried. 1 DSB between residues 214264.
C183 and C235 form disulphide
bond and are 30-40% SE. Ne more
buried. N, P and Hp more SE. Two
M 20-30% SE. most and more SE
are R res. E>D SE. H more buried.
W buried. His buried. 1 DSB
between residues 183-235.
P and Ne less SE. N and Hp more
SE. H buried. C194 and C274 40-50
and 30-40% SE respectively. 1 M
present which is buried. H158 90100 SE. W buried. More H buried.
Q< SE.
Short and
middle chain
fatty acids
(<C 12)
funnel-like
substratebinding sites
Organic
synthesis
Biodissel
-1.12-0.17
2 C surface exposed C190 and C270
which forms DSB. N and H more
SE. Ne more shielded than P. M255
Buried. H and D res are buried. E>D
SE. However D>E. W buried. All H
buried. Q< SE. 1 DSB between
residues 190-270.
Organic
synthesis
Unable to
predict
N and Hp more SE. D>E SE. N more
SE. C240 20-30% SE.
-
chloroform,
Most active in methanol and have 60% of
this activity in hexane.
-
PAL
Mesostable
Stable in n-hexane cyclohexan, p-xylene,
1-octanol, ethanol, pentanol inhibit it.
-
PLKW156
Thermostable
Neutral
-
Stable in >95% Acetone, methanol and
ethanol
Inhibited by anionic surfactants such
as SDS and LBS (sodium
laurylbenzenesulfonate).
PCL
thermostable
Neutral
Stable in methanol, ethanol, acetone,
acetonitrile, DMF, DMSO and dioxane
Benzene or n-hexane had inhibitory effect
RLM
Thermostable
Alkalophilic
Stable in tert-butanol, hexane
dichloromethane, ethyl acetate, ethanolamine
3
-
C182 40-50% SE. All H and M
buried. Ne more SE than P at 6070% & 10-20%. P more in 10-30%.
More L SE. W buried. Q SE
1 DSB between residues 182-240.
In DMSO and methanol showed no activity.
PS5L
Thermostable
Alkalophilic
Acetonitrile and 1-butanol inhibitory
DMFA, acetone, ethanol, and isopropanol
decreased activity
n-hexane, tween -60, tween-80
-
Subfamily 4
GthLYN
Temperature
stability
Thermostable
Organic
synthesis,
detergent
>2
Max C SE. H res very less those
present are SE. R>K SE. E res more
SE. Out of 12 C 8 are SE. P>Ne Hp
and N are equivalently distributed.
Maximum Y buried. W SE. F more
buried. H SE.
pH stability
Organic stability
Substrate
Application
Log P
*Surface properties
Alkalophilic
Stable in 1%sds edta and
mercaptoethanol, ethanol.
Medium chain
length
triglycerides
Biodissel,
organic
synthesis
Unable to predict
Frequency of Hp and N > SE. P>
Ne SE. H140 and H312 50-60%
SE. M351 10-20% SE. H153 1020% SE. H85 and H152 20-30%
SE. 5 M and 2 C res buried. R> K
SE. H res equivalently SE and
buried. W SE.Y more SE. L more
buried.
C296 and C65 buried. M174 1020% SE. 6 His SE, 7 buried. 4 M
buried. > number of D SE but E
more SE than D. P more SE. Hp
and N equal contribution at 70-90
and 40-50%, At 20-30% >Hp res.
W SE. Y & F> SE.
P > SE. > N and Hp than P and Ne.
M173 10-20% SE. 5 M and 2 C
buried. 6 H SE and 6 buried. W
shows equivalent distribution. Y >
SE.
DTT, DMSO and isopropanol showed
slight inhibition
BTL
do
do
Cholate, CHAPS, ethanol for 1 hour
at 30 degree then decreases
Medium chain
length
triglycerides
Detergent,
environment
>2<4
BL42
do
do
Stable benzene hexane, decanol,
isooctane, tetradecane water
immiscible solvents enhanced
activity.
Medium chain
length
triglycerides
low
phospholipase
activity
Detergent,
organic
synthesis,
biodissel
>2
More stable in >4
Polar solvents reduced activity
4
BSLP1
do
do
Chaps and Triton X-100
Short-Medium
chain length
triglycerides
Detergent,
environment
<4
Medium chain
length
triglycerides
Dertergent,
Environmen
t
>2
Organic
synthesis
>2
Organic
synthesis,
detergent,
environment
>2
N res moreSE. P more Se. More D
SE. R >K in SE. H312 60-70% SE,
H140 and H223 50-60% SE. H153
40-50% SE, H85 and H152 2030% SE. 2 M 10-20% SE. 2 C
buried. 8 H and 3 M buried. W
buried. >F buried. >Y SE. > L
buried.
Maximum N SE. P> SE. Ne more
in 70-80%, 60-70% and 10-20%.
M173 10-20% SE. H223, H140,
H152, 153, 85, 87 SE. 5 H res
buried, C64 and C295 buried. 5 M
buried. D>E SE. > W SE. > L
buried. > M buried.
C98 80-90% SE. C22 40-50% SE.
C25 30-40% SE. C343 10-20%
SE. 3 C buried. 2 M buried. 4 H
SE and 8 H buried. D more SE
than E. K more SE than R. N>Hp
SE. W > buried. Y> SE.
P >Ne SE. N more SE. K > R.
H274, H146, H300, H143, H158,
H55 are 70-80, 60-70, 50-60, 3040 and 10-20% SE. M256 50-60%,
M104, M256 10-20% SE. 6 M
buried. E more SE. > Y SE. W
SE. > G SE.
SDS, Tween20 inhibitted
GLTW1
do
do
DTT, SDS, 1%detergents (Tween 20,
Chaps or Triton X-100)
SSL
-
-
p-xylene, benzene, toluene and
hexane
-
SAL3
Thermostable
Alkalophilic
20mM Triton and TX-100, NaDC,
NaTDC, and Tween 80, H2O2,
Sodium
perborate.
Short-chain fatty
acid esters
Subfamily 5
Temperature
stability
pH stability
Organic stability
Substrate
Application
Log P
*Surface properties
BSL
Mesostable
Alkalophilic
1-butanol, hexanol, benzene
and toluene
Short chain
triglycerides
Biodissel
Organic synthesis
Around
2
BLL
Thermostable
Alkalophilic
Various detergents and
organic solvents
Short chain
triglycerides
Varied
P more SE. N res more SE. C absent.
Hp more in 50-60%. M134, M137
and M78 SE. H152, H3, H156 SE. 1
M and 2 H. res buried. K>SE. D more
and > SE. W SE. Y> SE.
N res > SE. P res more in 60-70% and
10-30%. No C present. 3 M SE. 2 M
buried. >K SE. R more SE than K.
5
-
H184, H35 H188 SE. 2 H res buried.
W SE
Subfamily 6
PLB-11
BSL205y
Temperature
stability
Mesostable
pH stability
Organic stability
Subtrate
Application
LogP
*Surface properties
Alkalophilic
Short chain
triglycerides
Detergent,
biodissel
-1.12-0.33
Moderate
thermostable
Neutral
Ethanol, Methanol,
Dimethyl sulphoxide,
Dimethyl formamide
p-xylene, n-decane
Short chain
triglycerides
Organic
synthesis
>2-4
<4
3 C and 1 M buried. C136 30-40% SE. C79 20-30% SE. C221
10-20% SE. Ne res more SE than P. N res more SE. E > D SE.
>W SE. >H SE. >L buried.> M SE.
H206 50-60 SE. H104 30-40% SE. H 340 and M81 10-20%
SE. C absent. 6 M buried. 3 H res buried. N res more SE. Hp
less SE. Nothing conclusive can be said about P and Ne
residues. > W buried. 7 L SE, 11 buried. >F SE. >M buried.
Complete deactivation
in acetonitrile
No activity in
hexadecane
Subfamily 7
TTL
Temperature
stability
Thermostable
pH stability
Organic stability
Substrate
Application
LogP
*Surface properties
Alkalophilic
tert-butanol,
acetonitrile,
isopropanol, pyridine,
acetone up to a
concentration of 99%
(v/v) , DTT
(dithiotreitol),
guanidine
hydrochloride, pHMB
(phydroxymercuribenz
oate) and iodo-acetate
Medium chain
length
triglycerides
Varied
-0.340.77
No C. Ne> P SE. K more SE and > R. H19 10-20%,
H233 20-30%, H169 50-60%, H45 70-80%, SE. 6 M
buried. M147 60-70%, M1 50-60%, M158 20-30%,
M114 10-20 , M17 SE. W more buried. L<SE. H> SE.
Y>SE absent in core. G>SE. Q very less and least SE.
6
Subfamily 8
SRL
Subfamily 9
GGYO5l
Subfamily 10
SML
Temperature
stability
Thermostable
pH stability
Temperature
stability
Moderate
thermostable
pH stability
Temperature
stability
Moderate
thermostable
pH stability
Alkalophilic
Neutralalkaline
Neutralalkaline
Organic
stability
Water
miscible
organic
solvents,
acetone,
methanol,
ethanol,
isopropanol,
DMSO
Substrate
Application
Log P
*Surface properties
-
Biodissel,
detergent,
organic
synthesis
>2
Ne > SE. Hp < SE. Ne>P. C288 20-30% SE.
C276 10-20% SE. M518 20-30% SE. D>E SE.
M523 40-50% SE. H370, H491 40-50%H298,
H453, H243, H41, H492 20-30%, H188 1020% SE. 5 M and 2 C buried. R are buried but
K SE. W<SE. F<SE. L, A, V more buried.
Organic
stability
Methanol,
glycerol,
hexane, nheptane,
cyclohexane,
isooctane,
benzene, and
diethyl ether,
DMSO
Substrate
Application
Log P
*Surface properties
-
Biodissel,
detergent,
organic
synthesis
>2
Ne > SE. Hp < SE. Ne>P. C288 20-30% SE.
C276 10-20% SE. M518 20-30% SE. D>E SE.
M523 40-50% SE. H370, H491 40-50%H298,
H453, H243, H41, H492 20-30%, H188 1020% SE. 5 M and 2 C buried. R are buried but
K SE. W<SE. F<SE. L, A, V more buried.
Organic
stability
Various water
miscible and
immiscible
Substrate
Application
Log P
*Surface properties
Varied
<2
N > SE. Ne>SE than P. Hp res less SE. K>SE.
M1 30-40% and M324 10-20% SE. 1 M
buried. C absent D> SE than E. W maximum
SE. F<SE.
-
Hp-hydrophobic, N- neutral, P-positive, Ne-negative residues, DSB- Disulphide bond, SE- Surface exposed and res-residues. Amino acid is represented by their single letter codes * Surface
properties were analyzed through ASAview server. – represents unknown.
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