Table 1.

The function of residues in CGTase, according to B. circulans strain 251 numbering

Positions	Residue	Mutations	Function/effect
Acceptor subsite +2
183	Phe, Trp	Asn, Ser, Leu	Cyclization, reduction of hydrolysisa,b,c,d
232	Lys, Ala	Leu, Glu	General activityc,e
259	Phe, Tyr	Leu, Asn, Ile, Glu, His, Arg	Cyclization, reduction of hydrolysisa,b,c,d,f,g
Acceptor subsite +1
194	Leu	Thr	Cyclizationh
195	Tyr, Phe	All possibilities	Cyclizationa,b,f,h,i,j
230	Ala	Val	Transglycosylation specificityd,e,k
233	His	Asp, Asn, Arg	General activityl,m
Catalytic subsite −1
100	Tyr	Phe, Ser	General activityn,o
140	His	Asp, Asn, Arg	General activityl,m
227	Arg	Ala, Lys	General activityp
229	Asp	Ala, Asn	Catalytic nucleophileq,r,s,t
257	Glu	Ala, Gln	Acid–base catalystq,s,t
327	His	Asp, Asn, Arg	General activityl,m
328	Asp	Asn, Ala	General activity, transition state stabilisers,t
Donor subsite –2
98	His	Asp	General activitym
Donor subsite −3
47	Arg, Lys, His, Thr	Trp, Leu, Gln	Cyclizationk,u
89	Tyr, Asp, Ser, Gln, Ala, Gly	Phe, Ser, Asp, Gly	General activity, product specificityo,v
94	Asn, Ser, Tyr, Glu	Ser, Gln	Slightly decreased activitieso,k
196	Asp, Asn, Gly	His	Reduced activity, more α-cyclodextrinw
371	Asp	Gly, Asn, Arg	General activity, less α-cyclodextrinw
Donor subsites −4/−5	No side-chain interactions
Donor subsite –6
167	Tyr	Phe	Selection of long substratesx
179	Gly	Leu	Selection of long substratesx
180	Gly, Asn	Leu	Selection of long substratesx
193	Asn	Leu, Gly	Selection of long substrates x
Donor subsite –7
145	Ser, Met, Leu, Asp, Glu	Ala, Glu	Cyclodextrin specificity
146	Ser, Glu, Val, Phe, Leu	Pro	Cyclodextrin specificityv
147	Thr, Asp, Asn	Trp	Cyclodextrin specificity
Loop 145–151	See Fig. 3	Asp	Cyclodextrin specificityh
Other important residues
21	Phe, Val	Leu	Limitation of hydrolysisk
76	Ser, Thr	Pro	Hydrolysisn
135	Asp	Asn, Ala	Influences conformation of Glu257 and Arg227p
326	Asp, Ser	Asp, Gln, Leu	Cyclization and general activityw

^a(Leemhuis et al. 2002a)

^b(Nakamura et al. 1994)

^c(van der Veen et al. 2001)

^d(Kelly et al. 2007)

^e(Kelly et al. 2008a)

^f(Fujiwara et al. 1992a)

^g(Shim et al. 2004)

^h(Parsiegla et al. 1998)

ⁱ(Penninga et al. 1995)

^j(Sin et al. 1994)

^k(Leemhuis et al. 2003d)

^l(Nakamura et al. 1993)

^m(Mattsson et al. 1995)

ⁿ(Kelly et al. 2008b)

^o(Kim et al. 1997)

^p(Leemhuis et al. 2003c)

^q(Uitdehaag et al. 1999b)

^r(Mosi et al. 1997)

^s(Knegtel et al. 1995)

^t(Klein et al. 1992)

^u(van der Veen et al. 2000a)

^v(van der Veen et al. 2000b)

^w(Wind et al. 1998)

^x(Leemhuis et al. 2002b)